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  • 1.
    Adler, Jeremy
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology.
    Parmryd, Ingela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Quantification of Colocalisation; Co-Occurrence, Correlation, Empty Voxels, Regions of Interest and Thresholding2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 602A-602A p.Article in journal (Other academic)
    Abstract [en]

    Measuring colocalisation is not straightforward with a plethora of coefficients that encapsulate different definitions. Measurements may also be implemented differently. Not only do measurements differ; interconversion is impossible making comparisons challenging. There is a need to cull coefficients and for clear definitions of what precisely is meant by colocalisation in individual studies. Colocalisation can be considered to have two components; co-occurrence which reports whether the fluorophores are found together and correlation which reports on the similarity in their patterns of intensity.

  • 2. Alizadehheidari, Mohammadreza
    et al.
    Werner, Erik
    Noble, Charleston
    Nyberg, Lena
    Fritzsche, Joachim
    Mehlig, Bernhard
    Tegenfeldt, Jonas
    Ambjoernsson, Tobias
    Persson, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Westerlund, Fredrik
    Nanoconfined Circular DNA2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 274A-274A p.Article in journal (Other academic)
    Abstract [en]

    Nanofluidic channels have become a versatile tool to manipulate single DNA molecules. They allow investigation of confined single DNA molecules from a fundamental polymer physics perspective as well as for example in DNA barcoding techniques.

  • 3. Allahverdiyeva, Yagut
    et al.
    Mamedov, Fikret
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström.
    Mäenpää, Pirkko
    Vass, Imre
    Aro, Eva-Mari
    Modulation of photosynthetic electron transport in the absence of terminal electron acceptors: characterization of the rbcL deletion mutant of tobacco2005In: Biochimica et Biophysica Acta (BBA) - Bioenergetics, ISSN 0005-2728, Vol. 1709, no 1, 69-83 p.Article in journal (Refereed)
  • 4.
    Almaqwashi, Ali A.
    et al.
    Northeastern Univ, Dept Phys, Boston, MA 02115 USA..
    Andersson, Johanna
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC. Chalmers, Dept Chem & Chem Engn, S-41296 Gothenburg, Sweden..
    Lincoln, Per
    Chalmers, Dept Chem & Chem Engn, S-41296 Gothenburg, Sweden..
    Rouzina, Ioulia
    Ohio State Univ, Dept Chem & Biochem, Columbus, OH 43210 USA..
    Westerlund, Fredrik
    Chalmers, Dept Biol & Biol Engn, S-41296 Gothenburg, Sweden..
    Williams, Mark C.
    Northeastern Univ, Dept Phys, Boston, MA 02115 USA..
    Dissecting the Dynamic Pathways of Stereoselective DNA Threading Intercalation2016In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, no 6, 1255-1263 p.Article in journal (Refereed)
    Abstract [en]

    DNA intercalators that have high affinity and slow kinetics are developed for potential DNA-targeted therapeutics. Although many natural intercalators contain multiple chiral subunits, only intercalators with a single chiral unit have been quantitatively probed. Dumbbell-shaped DNA threading intercalators represent the next order of structural complexity relative to simple intercalators, and can provide significant insights into the stereoselectivity of DNA-ligand intercalation. We investigated DNA threading intercalation by binuclear ruthenium complex [mu-dppzip(phen)(4)Ru-2](4+) (Piz). Four Piz stereoisomers are defined by the chirality of the intercalating subunit (Ru(phen)(2)dppz) and the distal subunit (Ru(phen)(2)ip), respectively, each of which can be either right-handed (Delta) or left-handed (Lambda). We used optical tweezers to measure single DNA molecule elongation due to threading intercalation, revealing force-dependent DNA intercalation rates and equilibrium dissociation constants. The force spectroscopy analysis provided the zero-force DNA binding affinity, the equilibrium DNA-ligand elongation Delta x(eq), and the dynamic DNA structural deformations during ligand association x(on) and dissociation x(off). We found that Piz stereoisomers exhibit over 20-fold differences in DNA binding affinity, from a K-d of 27 +/- 3 nM for (Delta,Lambda)-Piz to a K-d of 622 +/- 55 nM for (Lambda,Delta)-Piz. The striking affinity decrease is correlated with increasing Delta x(eq) from 0.30 +/- 0.02 to 0.48 +/- 0.02 nm and x(on) from 0.25 +/- 0.01 to 0.46 +/- 0.02 nm, but limited x(off) changes. Notably, the affinity and threading kinetics is 10-fold enhanced for right-handed intercalating subunits, and 2- to 5-fold enhanced for left-handed distal subunits. These findings demonstrate sterically dispersed transition pathways and robust DNA structural recognition of chiral intercalators, which are critical for optimizing DNA binding affinity and kinetics.

  • 5.
    Amselem, Elias
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Marklund, Emil
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Kipper, Kalle
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Johansson, Magnus
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Deindl, Sebastian
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Elf, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Real- Time Single Protein Tracking with Polarization Readout using a Confocal Microscope2017In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, no 3, 295A-295A p.Article in journal (Other academic)
  • 6. Aquila, Andrew
    et al.
    Hunter, Mark S.
    Doak, R. Bruce
    Kirian, Richard A.
    Fromme, Petra
    White, Thomas A.
    Andreasson, Jakob
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Arnlund, David
    Bajt, Saša
    Barends, Thomas R. M.
    Barthelmess, Miriam
    Bogan, Michael J.
    Bostedt, Christoph
    Bottin, Hervé
    Bozek, John D.
    Caleman, Carl
    Coppola, Nicola
    Davidsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Physical Chemistry.
    DePonte, Daniel P.
    Elser, Veit
    Epp, Sascha W.
    Erk, Benjamin
    Fleckenstein, Holger
    Foucar, Lutz
    Frank, Matthias
    Fromme, Raimund
    Graafsma, Heinz
    Grotjohann, Ingo
    Gumprecht, Lars
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hampton, Christina Y.
    Hartmann, Andreas
    Hartmann, Robert
    Hau-Riege, Stefan
    Hauser, Günter
    Hirsemann, Helmut
    Holl, Peter
    Holton, James M.
    Hömke, André
    Johansson, Linda
    Kimmel, Nils
    Kassemeyer, Stephan
    Krasniqi, Faton
    Kühnel, Kai-Uwe
    Liang, Mengning
    Lomb, Lukas
    Malmerberg, Erik
    Marchesini, Stefano
    Martin, Andrew V.
    Maia, Filipe R.N.C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Messerschmidt, Marc
    Nass, Karol
    Reich, Christian
    Neutze, Richard
    Rolles, Daniel
    Rudek, Benedikt
    Rudenko, Artem
    Schlichting, Ilme
    Schmidt, Carlo
    Schmidt, Kevin E.
    Schulz, Joachim
    Seibert, M. Marvin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Soltau, Heike
    Shoeman, Robert L.
    Sierra, Raymond
    Starodub, Dmitri
    Stellato, Francesco
    Stern, Stephan
    Strüder, Lothar
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Ullrich, Joachim
    Wang, Xiaoyu
    Williams, Garth J.
    Weidenspointner, Georg
    Weierstall, Uwe
    Wunderer, Cornelia
    Barty, Anton
    Spence, John C. H.
    Chapman, Henry N.
    Time-resolved protein nanocrystallography using an X-ray free-electron laser2012In: Optics Express, ISSN 1094-4087, E-ISSN 1094-4087, Vol. 20, no 3, 2706-2716 p.Article in journal (Refereed)
    Abstract [en]

    We demonstrate the use of an X-ray free electron laser synchronized with an optical pump laser to obtain X-ray diffraction snapshots from the photoactivated states of large membrane protein complexes in the form of nanocrystals flowing in a liquid jet. Light-induced changes of Photosystem I-Ferredoxin co-crystals were observed at time delays of 5 to 10 µs after excitation. The result correlates with the microsecond kinetics of electron transfer from Photosystem I to ferredoxin. The undocking process that follows the electron transfer leads to large rearrangements in the crystals that will terminally lead to the disintegration of the crystals. We describe the experimental setup and obtain the first time-resolved femtosecond serial X-ray crystallography results from an irreversible photo-chemical reaction at the Linac Coherent Light Source. This technique opens the door to time-resolved structural studies of reaction dynamics in biological systems.

  • 7.
    Ashrafzadeh, Parham
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Dinic, Jelena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Parmryd, Ingela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Actin Filaments Attachment to the Plasma Membrane Cause the Formation of Ordered Lipid Domains in Live Cells2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 706A-706A p.Article in journal (Other academic)
    Abstract [en]

    The aim of this study was to investigate the relationship between ordered plasma membrane nanodomains and actin filaments using di-4-ANEPPDHQ and laurdan together with the reagents that affect actin filament dynamics in live Jurkat and primary T cells. The degree of lipid packing can be quantified using polarity sensitive membrane dyes such as laurdan and di-4-ANEPPDHQ. These two dyes display a red shift in their emission peaks for membranes in ld phase relative to lo phase. Laurdan is uncharged and can easily flip between two leaflets of the plasma membrane and we demonstrate that it reports equally on the two leaflets of the plasma membrane.

  • 8.
    Ashrafzadeh, Parham
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Parmryd, Ingela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Methods applicable to membrane nanodomain studies?2015In: Essays in Biochemistry, ISSN 0071-1365, E-ISSN 1744-1358, Vol. 57, 57-68 p.Article, review/survey (Refereed)
    Abstract [en]

    Membrane nanodomains are dynamic liquid entities surrounded by another type of dynamic liquid. Diffusion can take place inside, around and in and out of the domains, and membrane components therefore continuously shift between domains and their surroundings. In the plasma membrane, there is the further complexity of links between membrane lipids and proteins both to the extracellular matrix and to intracellular proteins such as actin filaments. In addition, new membrane components are continuously delivered and old ones removed. On top of this, cells move. Taking all of this into account imposes great methodological challenges, and in the present chapter we discuss some methods that are currently used for membrane nanodomain studies, what information they can provide and their weaknesses.

  • 9.
    Balzarotti, Francisco
    et al.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Eilers, Yvan
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Gwosch, Klaus C.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Gynnå, Arvid H.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Westphal, Volker
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany..
    Stefani, Fernando D.
    Consejo Nacl Invest Cient & Tecn, Ctr Invest Bionanociencias CIBION, Buenos Aires, DF, Argentina.;Univ Buenos Aires, Fac Ciencias Exactas & Nat, Dept Fis, Buenos Aires, DF, Argentina..
    Elf, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Hell, Stefan W.
    Max Planck Inst Biophys Chem, Dept NanoBiophoton, Gottingen, Germany.;Max Planck Inst Med Res, Dept Opt Nanoscopy, Heidelberg, Germany.;German Canc Res Ctr, Opt Nanoscopy Div, Heidelberg, Germany..
    Nanometer resolution imaging and tracking of fluorescent molecules with minimal photon fluxes2017In: Science, ISSN 0036-8075, E-ISSN 1095-9203, Vol. 355, no 6325, 606-612 p.Article in journal (Refereed)
    Abstract [en]

    We introduce MINFLUX, a concept for localizing photon emitters in space. By probing the emitter with a local intensity minimum of excitation light, MINFLUX minimizes the fluorescence photons needed for high localization precision. In our experiments, 22 times fewer fluorescence photons are required as compared to popular centroid localization. In superresolutionmicroscopy, MINFLUXattained similar to 1-nanometer precision, resolving molecules only 6 nanometers apart. MINFLUX tracking of single fluorescent proteins increased the temporal resolution and the number of localizations per trace by a factor of 100, as demonstrated with diffusing 30S ribosomal subunits in living Escherichia coli. As conceptual limits have not been reached, we expect this localization modality to break new ground for observing the dynamics, distribution, and structure of macromolecules in living cells and beyond.

  • 10.
    Barrozo, Alexandre H.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Carvalho, Alexandra Pires
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Understanding Functional Evolution in the Alkaline Phosphatase Superfamily2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 675A-675A p.Article in journal (Other academic)
    Abstract [en]

    Over the past 40 years, it has been demonstrated that many enzymes are capable of promiscuous catalytic activities, facilitating the turnover of more than one chemically distinct substrate. This has been argued to play an important role in enzyme evolution, with highly promiscuous progenitor enzymes evolving under evolutionary pressure to modern day specialists, while still retaining some level of their former promiscuous activities1. This theory has been extensively tested by different experiments using in vitro evolution2. The alkaline phosphatase superfamily members provide a particularly attractive showcase for studying enzyme promiscuity, as they often show reciprocal promiscuity, in that the native reaction for one member is often a side-reaction for another3. While deceptively similar, their catalyzed reactions (cleavage of P-O and S-O bonds) proceed via distinct transition states and protonation requirements4,5. We present detailed computational studies of the promiscuous catalytic activity of three evolutionarily related members: the arylsulfatase from Pseudomonas aeruginosa6, and the phosphonate monoester hydrolases from Burkholderia caryophili7and Rhizobium leguminosarum8. By tracking their structural and electrostatic features, and comparing to other known members of the superfamily, we provide an atomic-level map for functional evolution within this superfamily.

  • 11. Bauer, Pavol
    et al.
    Engblom, Stefan
    Mikulovic, Sanja
    Senek, Aleksandar
    Multiscale modeling via split-step methods in neural firingIn: Mathematical and Computer Modelling of Dynamical Systems, ISSN 1387-3954, E-ISSN 1744-5051Article in journal (Refereed)
    Abstract [en]

    Neuronal models based on the Hodgkin-Huxley equation form a  fundamental framework in the field of computational  neuroscience. While the neuronal state is often modeled  deterministically, experimental recordings show stochastic  fluctuations, presumably driven by molecular noise from the  underlying microphysical conditions. In turn, the firing of  individual neurons gives rise to an electric field in extracellular  space, also thought to affect the firing pattern of nearby neurons.  We develop a multiscale model which combines a stochastic ion  channel gating process taking place on the neuronal membrane,  together with the propagation of an action potential along the  neuronal structure. We also devise a numerical method relying on a  split-step strategy which effectively couples these two processes  and we experimentally test the feasibility of this approach. We  finally also explain how the approach can be extended with Maxwell's  equations to allow the potential to be propagated in extracellular  space.

  • 12.
    Berg, Otto G.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Mahmutovic, Anel
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Marklund, Emil
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    Elf, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Systems Biology.
    The helical structure of DNA facilitates binding2016In: Journal of Physics A: Mathematical and Theoretical, ISSN 1751-8113, E-ISSN 1751-8121, Vol. 9, no 36, 364002Article in journal (Other academic)
    Abstract [en]

    The helical structure of DNA imposes constraints on the rate of diffusion-limited protein binding. Here we solve the reaction-diffusion equations for DNA-like geometries and extend with simulations when necessary. We find that the helical structure can make binding to the DNA more than twice as fast compared to a case where DNA would be reactive only along one side. We also find that this rate advantage remains when the contributions from steric constraints and rotational diffusion of the DNA-binding protein are included. Furthermore, we find that the association rate is insensitive to changes in the steric constraints on the DNA in the helix geometry, while it is much more dependent on the steric constraints on the DNA-binding protein. We conclude that the helical structure of DNA facilitates the nonspecific binding of transcription factors and structural DNA-binding proteins in general.

  • 13.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology.
    Protein Crystallization: Second Edition2009 (ed. 2)Book (Other academic)
  • 14.
    Bergh, Magnus
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Interaction of Ultrashort X-ray Pulses with Material2007Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Radiation damage limits the resolution in imaging experiments. Damage is caused by energy deposited into the sample during exposure. Ultrashort and extremely bright X-ray pulses from free-electron lasers (FELs) offer the possibility to outrun key damage processes, and temporarily improve radiation tolerance. Theoretical models indicate that high detail-resolutions could be realized on non-crystalline samples with very short pulses, before plasma expansion.

    Studies presented here describe the interaction of a very intense and ultrashort X-ray pulse with material, and investigate boundary conditions for flash diffractive imaging both theoretically and experimentally. In the hard X-ray regime, predictions are based on particle simulations with a continuum formulation that accounts for screening from free electrons.

    First experimental results from the first soft X-ray free-electron laser, the FLASH facility in Hamburg, confirm the principle of flash imaging, and provide the first validation of our theoretical models. Specifically, experiments on nano-fabricated test objects show that an interpretable image can be obtained to high resolution before the sample is vaporized. Radiation intensity in these experiments reached 10^14 W/cm^2, and the temperature of the sample rose to 60000 Kelvin after the 25 femtosecond pulse left the sample. Further experiments with time-delay X-ray holography follow the explosion dynamics over some picoseconds after illumination.

    Finally, this thesis presents results from biological flash-imaging studies on living cells. The model is based on plasma calculations and fluid-like motions of the sample, supported by the time-delay measurements. This study provides an estimate for the achievable resolutions as function of wavelength and pulse length. The technique was demonstrated by our team in an experiment where living cells were exposed to a single shot from the FLASH soft X-ray laser.

    List of papers
    1. Model for the Dynamics of a Water Cluster in an X-ray Free Electron Laser Beam
    Open this publication in new window or tab >>Model for the Dynamics of a Water Cluster in an X-ray Free Electron Laser Beam
    2004 (English)In: Physical Review E. Statistical, Nonlinear, and Soft Matter Physics, ISSN 1539-3755, Vol. 70, no 5:1, 051904- p.Article in journal (Refereed) Published
    Abstract [en]

    A microscopic sample placed into a focused x-ray free electron laser beam will explode due to strong ionization on a femtosecond time scale. The dynamics of this Coulomb explosion has been modeled by Neutze et al. [Nature (London) 406, 752 (2000)] for a protein, using computer simulations. The results suggest that by using ultrashort exposures, structural information may be collected before the sample is destroyed due to radiation damage. In this paper a method is presented to include the effect of screening by free electrons in the sample in a molecular dynamics simulation. The electrons are approximated by a classical gas, and the electron distribution is calculated iteratively from the Poisson-Boltzmann equation. Test simulations of water clusters reveal the details of the explosion dynamics, as well as the evolution of the free electron gas during the beam exposure. We find that inclusion of the electron gas in the model slows down the Coulomb explosion. The hydrogen atoms leave the sample faster than the oxygen atoms, leading to a double layer of positive ions. A considerable electron density is located between these two layers. The fact that the hydrogens are found to explode much faster than the oxygens means that the diffracting part of the sample stays intact somewhat longer than the sample as a whole.

    Keyword
    Computer Simulation, Electrons, Lasers, Models
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:uu:diva-96323 (URN)15600653 (PubMedID)
    Available from: 2007-10-24 Created: 2007-10-24 Last updated: 2013-02-26Bibliographically approved
    2. Soft-x-ray free-electron-laser interaction with materials
    Open this publication in new window or tab >>Soft-x-ray free-electron-laser interaction with materials
    2007 (English)In: Physical Review E. Statistical, Nonlinear, and Soft Matter Physics, ISSN 1063-651X, Vol. 76, no 4, 046403- p.Article in journal (Refereed) Published
    Abstract [en]

    Soft-x-ray free-electron lasers have enabled materials studies in which structural information is obtained faster than the relevant probe-induced damage mechanisms. We present a continuum model to describe the damage process based on hot-dense plasma theory, which includes a description of the energy deposition in the samples, the subsequent dynamics of the sample, and the detector signal. We compared the model predictions with experimental data and mostly found reasonable agreement. In view of future free-electron-laser performance, the model was also used to predict damage dynamics of samples and optical elements at shorter wavelengths and larger photon fluences than currently available.

    National Category
    Physical Sciences
    Identifiers
    urn:nbn:se:uu:diva-96324 (URN)10.1103/PhysRevE.76.046403 (DOI)000250622100073 ()
    Available from: 2007-10-24 Created: 2007-10-24 Last updated: 2010-02-19Bibliographically approved
    3. Force Field Benchmark of Organic Liquids: Density, Enthalpy of Vaporization, Heat Capacities, Surface Tension, Isothermal Compressibility, Volumetric Expansion Coefficient, and Dielectric Constant
    Open this publication in new window or tab >>Force Field Benchmark of Organic Liquids: Density, Enthalpy of Vaporization, Heat Capacities, Surface Tension, Isothermal Compressibility, Volumetric Expansion Coefficient, and Dielectric Constant
    Show others...
    2007 (English)In: Physical Review Letters, ISSN 0031-9007, Vol. 98, no 14, 145502- p.Article in journal (Refereed) Published
    Abstract [en]

    At the recently built FLASH x-ray free-electron laser, we studied the reflectivity of Si/C multilayers with fluxes up to 3×1014W/cm2. Even though the nanostructures were ultimately completely destroyed, we found that they maintained their integrity and reflectance characteristics during the 25-fs-long pulse, with no evidence for any structural changes over lengths greater than 3Å. This experiment demonstrates that with intense ultrafast pulses, structural damage does not occur during the pulse, giving credence to the concept of diffraction imaging of single macromolecules.

    Keyword
    X-ray effects, Optical properties of low-dimensional, mesoscopic, and nanoscale materials and structures
    National Category
    Biological Sciences Physical Sciences
    Identifiers
    urn:nbn:se:uu:diva-96325 (URN)10.1103/PhysRevLett.98.145502 (DOI)000245512100037 ()
    Note


    Available from: 2007-10-24 Created: 2007-10-24 Last updated: 2016-04-12Bibliographically approved
    4. Feasibility of imaging living cells at subnanometer resolutions by ultrafast X-ray diffraction
    Open this publication in new window or tab >>Feasibility of imaging living cells at subnanometer resolutions by ultrafast X-ray diffraction
    Show others...
    2008 (English)In: Quarterly reviews of biophysics (Print), ISSN 0033-5835, E-ISSN 1469-8994, Vol. 41, no 3-4, 181-204 p.Article, review/survey (Refereed) Published
    Abstract [en]

    Detailed structural investigations on living cells are problematic because existing structural methods cannot reach high resolutions on non-reproducible objects. Illumination with an ultrashort and extremely bright X-ray pulse can outrun key damage processes over a very short period. This can be exploited to extend the diffraction signal to the highest possible resolution in flash diffraction experiments. Here we present an analysis or the interaction of a very intense and very short X-ray pulse with a living cell, using a non-equilibrium population kinetics plasma code with radiation transfer. Each element in the evolving plasma is modeled by numerous states to monitor changes in the atomic populations as a function of pulse length, wavelength, and fluence. The model treats photoionization, impact ionization, Auger decay, recombination, and inverse bremsstrahlung by solving rate equations in a self-consistent manner and describes hydrodynamic expansion through the ion sound speed, The results show that subnanometer resolutions could be reached on micron-sized cells in a diffraction-limited geometry at wavelengths between 0.75 and 1.5 nm and at fluences of 10(11)-10(12) photonS mu M (2) in less than 10 fs. Subnanometer resolutions could also be achieved with harder X-rays at higher fluences. We discuss experimental and computational strategies to obtain depth information about the object in flash diffraction experiments.

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:uu:diva-96326 (URN)10.1017/S003358350800471X (DOI)000262098500001 ()
    Available from: 2007-10-24 Created: 2007-10-24 Last updated: 2013-03-01Bibliographically approved
    5. Interaction of Ultrashort X-ray Pulses with B4C, SiC and Si
    Open this publication in new window or tab >>Interaction of Ultrashort X-ray Pulses with B4C, SiC and Si
    2008 (English)In: Physical Review E. Statistical, Nonlinear, and Soft Matter Physics: Statistical Physics, Plasmas, Fluids, and Related Interdisciplinary Topics, ISSN 1063-651X, E-ISSN 1095-3787, Vol. 77, no 2, 026404-1-026404-8 p.Article in journal (Refereed) Published
    Abstract [en]

    The interaction of 32.5 and 6 nm ultrashort x-ray pulses with the solid materials B4C, SiC, and Si is simulated with a nonlocal thermodynamic equilibrium radiation transfer code. We study the ionization dynamics as a function of depth in the material and modifications of the opacity during irradiation, and estimate the crater depth. Furthermore, we compare the estimated crater depth with experimental data, for fluences up to 2.2 J/cm(2). Our results show that, at 32.5 nm irradiation, the opacity changes by less than a factor of 2 for B4C and Si and by a factor of 3 for SiC, for fluences up to 200 J/cm(2). At a laser wavelength of 6 nm, the model predicts a dramatic decrease in opacity due to the weak inverse bremsstrahlung, increasing the crater depth for high fluences.

    Keyword
    boron compounds, bremsstrahlung, elemental semiconductors, high-speed optical techniques, ionisation, laser beam effects, opacity, silicon, silicon compounds, thermodynamics, wide band gap semiconductors, X-ray effects
    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-96327 (URN)10.1103/PhysRevE.77.026404 (DOI)000253763800053 ()18352130 (PubMedID)
    Available from: 2007-10-24 Created: 2007-10-24 Last updated: 2013-02-27Bibliographically approved
  • 15.
    Blokzijl, Andries
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
    Nong, Rachel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
    Darmanis, S.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Hertz, E.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Landegren, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
    Kamali-Moghaddam, Masood
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
    Protein biomarker validation via proximity ligation assays2014In: Biochimica et Biophysica Acta - Proteins and Proteomics, ISSN 1570-9639, E-ISSN 1878-1454, Vol. 1844, no 5, 933-939 p.Article, review/survey (Refereed)
    Abstract [en]

    The ability to detect minute amounts of specific proteins or protein modifications in blood as biomarkers for a plethora of human pathological conditions holds great promise for future medicine. Despite a large number of plausible candidate protein biomarkers published annually, the translation to clinical use is impeded by factors such as the required size of the initial studies, and limitations of the technologies used. The proximity ligation assay (PLA) is a versatile molecular tool that has the potential to address some obstacles, both in validation of biomarkers previously discovered using other techniques, and for future routine clinical diagnostic needs. The enhanced specificity of PIA extends the opportunities for large-scale, high-performance analyses of proteins. Besides advantages in the form of minimal sample consumption and an extended dynamic range, the PLA technique allows flexible assay reconfiguration. The technology can be adapted for detecting protein complexes, proximity between proteins in extracellular vesicles or in circulating tumor cells, and to address multiple post-translational modifications in the same protein molecule. We discuss herein requirements for biomarker validation, and how PLA may play an increasing role in this regard. We describe some recent developments of the technology, including proximity extension assays, the use of recombinant affinity reagents suitable for use in proximity assays, and the potential for single cell proteomics. This article is part of a Special Issue entitled: Biomarkers: A Proteomic Challenge. (C) 2013 Elsevier B.V. All rights reserved.

  • 16.
    Cai, Yixiao
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Applied Materials Sciences.
    Lendel, Christofer
    Österlund, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Solid State Physics.
    Kasrayan, Alex
    Lannfelt, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Geriatrics.
    Ingelsson, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Geriatrics.
    Nikolajeff, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Applied Materials Sciences.
    Karlsson, Mikael
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Applied Materials Sciences.
    Bergström, Joakim
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Geriatrics.
    Changes in secondary structure of α-synuclein during oligomerization induced by reactive aldehydes.2015In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 464, no 1, 336-341 p.Article in journal (Refereed)
    Abstract [en]

    The oxidative stress-related reactive aldehydes 4-hydroxy-2-nonenal (HNE) and 4-oxo-2-nonenal (ONE) have been shown to promote formation of α-synuclein oligomers in vitro. However, the changes in secondary structure of α-synuclein and the kinetics of the oligomerization process are not known and were the focus of this study. Size exclusion chromatography showed that after 1 h of incubation, HNE induced the formation of an oligomeric α-synuclein peak with a molecular weight of about ∼2000 kDa, which coincided with a decreasing ∼50 kDa monomeric peak. With prolonged incubation (up to 24 h) the oligomeric peak became the dominating molecular species. In contrast, in the presence of ONE, a ∼2000 oligomeric peak was exclusively observed after 15 min of incubation and this peak remained constant with prolonged incubation. Western blot analysis of HNE-induced α-synuclein oligomers showed the presence of monomers (15 kDa), SDS-resistant low molecular (30-160 kDa) and high molecular weight oligomers (≥260 kDa), indicating that the oligomers consisted of both covalent and non-covalent protein. In contrast, ONE-induced α-synuclein oligomers only migrated as covalent cross-linked high molecular-weight material (≥300 kDa). Both circular dichroism (CD) and Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy showed that the formation of HNE- and ONE-induced oligomers coincided with a spectral change from random coil to β-sheet. However, ONE-induced α-synuclein oligomers exhibited a slightly higher degree of β-sheet. Taken together, our results indicate that both HNE and ONE induce a change from random coil to β-sheet structure that coincides with the formation of α-synuclein oligomers. Albeit through different kinetic pathways depending on the degree of cross-linking.

  • 17.
    Caleman, Carl
    et al.
    Physik Department E17, Technische Universität München.
    Ortiz, Carlos
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Materials Theory.
    Marklund, Erik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Bultmark, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Materials Theory.
    Gabrysch, Markus
    Uppsala University, Disciplinary Domain of Science and Technology, Technology, Department of Engineering Sciences, Electricity.
    Parak, F. G.
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Klintenberg, Mattias
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Materials Science, Materials Theory.
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Radiation damage in biological material: electronic properties and electron impact ionization in urea2009In: Europhysics letters, ISSN 0295-5075, E-ISSN 1286-4854, Vol. 85, no 1, 18005- p.Article in journal (Refereed)
    Abstract [en]

    Radiation damage is an unavoidable process when performing structural investigations of biological macromolecules with X-rays. In crystallography this process can be limited through damage distribution in a crystal, while for single molecular imaging it can be outrun by employing short intense pulses. Secondary electron generation is crucial during damage formation and we present a study of urea, as model for biomaterial. From first principles we calculate the band structure and energy loss function, and subsequently the inelastic electron cross-section in urea. Using Molecular Dynamics simulations, we quantify the damage and study the magnitude and spatial extent of the electron cloud coming from an incident electron, as well as the dependence with initial energy.

  • 18.
    Carlsson, Gunilla
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Crystallography in Four Dimensions: Methods and Applications2004Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The four-electron reduction of dioxygen to water is the most exothermic non-photochemical reaction available to biology. A detailed molecular description of this reaction is needed to understand oxygen-based redox processes. Horseradish peroxidase (HRP) is a haem-containing redox enzyme capable of catalysing the reduction of dioxygen to water. We developed instrumentation and experimental methodology to capture and characterise by X-ray crystallography transient reaction intermediates in this reaction.

    An instrument was designed (“the vapour stream system”) to facilitate reaction initiation, monitoring and intermediate trapping. In combination with single crystal microspectrophotometry, it was used to obtain conditions for capturing a reactive dioxygen complex in HRP. X-ray studies on oxidised intermediates can be difficult for various reasons. Electrons re-distributed in the sample through the photoelectric effect during X-ray exposure can react with high-valency intermediates. In order to control such side reactions during data collection, we developed a new method based on an angle-resolved spreading of the X-ray dose over many identical crystals. Composite data sets built up from small chunks of data represent crystal structures which received different X-ray doses. As the number of electrons liberated in the crystal is dose dependent, this method allows us to observe and drive redox reactions electron-by-electron in the crystal, using X-rays.

    The methods developed here were used to obtain a three-dimensional movie on the X-ray-driven reduction of dioxygen to water in HRP. Separate experiments established high resolution crystal structures for all intermediates, showing such structures with confirmed redox states for the first time.

    Activity of HRP is influenced by small molecule ligands, and we also determined the structures of HRP in complex with formate, acetate and carbon monoxide.

    Other studies established conditions for successfully trapping the M-intermediate in crystals of mutant bacteriorhodopsin, but the poor diffraction quality of these crystals prevented high-resolution structural studies.

    List of papers
    1. Protein crystallography in a vapour steam: data collection, reaction initiation and intermediate trapping in naked hydrated protein crystals
    Open this publication in new window or tab >>Protein crystallography in a vapour steam: data collection, reaction initiation and intermediate trapping in naked hydrated protein crystals
    Show others...
    2002 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 35, no 1, 113-116 p.Article in journal (Refereed) Published
    Abstract [en]

    A procedure is presented for experiments on naked unfrozen protein crystals with the crystal mounted in a conventional cryo-loop and surrounded by a stream of a wet gas. The composition and temperature of the vapour stream can be adjusted to keep the crystal without deterioration for many hours. The arrangement allows (i) for rapidly testing crystals for diffraction before freezing, (ii) for data collection between 268-303 K with greatly reduced background, (iii) for the controlled drying or wetting of crystals, (iv) for the anaerobic manipulation of protein crystals, and (v) for the introduction of gaseous or volatile ingredients and reactants into the crystal. The technique offers new experimental possibilities, e.g. in time-resolved structural studies. Reaction initiation in many protein crystals can be achieved by changing the composition of the vapour stream to create a new chemical environment around the crystal and to introduce substrates/reactants either in the gas phase or as microdroplets. Spectral changes during such reactions can be monitored by single-crystal microspectrophotometry, and, once an intermediate has been detected at high concentrations, the crystal can be frozen, e.g. by rapidly switching the warm vapour stream to a cryogenically cooled helium or nitrogen jet. Representative examples are presented in this paper.

    National Category
    Medical and Health Sciences
    Research subject
    Biochemistry
    Identifiers
    urn:nbn:se:uu:diva-91905 (URN)10.1107/S0021889801020702 (DOI)
    Available from: 2004-09-01 Created: 2004-09-01 Last updated: 2013-12-19Bibliographically approved
    2. Defining redox state of X-ray crystal structures by single-crystal ultraviolet visible microspectrophotometry
    Open this publication in new window or tab >>Defining redox state of X-ray crystal structures by single-crystal ultraviolet visible microspectrophotometry
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    2002 (English)In: Methods in Enzymology, ISSN 0076-6879, Vol. 353, 301-318 p.Article in journal (Refereed) Published
    Abstract [en]

    Exciting results have been emerging from the field of single-crystal X-ray crystallography, giving unprecedented detail of freeze-trapped reaction intermediates from important classes of macromolecules that contain chromophores. These structures have been coupled with single-crystal UV-visible microspectrophotometry. This has defined the distinct catalytic intermediates present in the crystal structures, allowing the correlation of electronic transitions with the observed structural transitions. Of particular note is that many of these structures have been generated “on the fly” during kinetic turnover in the crystal. Most enzymatic reactions proceed through distinct catalytic intermediates that, under favorable conditions, may accumulate transiently in the crystal during turnover. In some cases, the physical constraints of the contacts within crystals may also lead to a significant slowing of the reaction at certain points along the pathway where conformational changes are required. This can lead to a transient build-up of spectrally distinct intermediates in the crystal that can be trapped by flash freezing in liquid nitrogen, allowing a complete single-crystal data set to be collected to the highest possible resolution at a later time. Similar build-up of intermediates may be achieved by altering the pH, temperature, or the solvent environment around the protein in the crystal, or by producing engineered variants that build up an intermediate of interest. The chapter focuses on the technical considerations required to carry out UV-visible microspectroscopy of single crystals.

    National Category
    Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-91906 (URN)10.1016/S0076-6879(02)53057-3 (DOI)000176466500027 ()
    Available from: 2004-09-01 Created: 2004-09-01 Last updated: 2013-12-19Bibliographically approved
    3. The catalytic pathway of horseradish peroxidase at high resolution
    Open this publication in new window or tab >>The catalytic pathway of horseradish peroxidase at high resolution
    Show others...
    2002 In: Nature, Vol. 417, 463-468 p.Article in journal (Refereed) Published
    Identifiers
    urn:nbn:se:uu:diva-91907 (URN)
    Available from: 2004-09-01 Created: 2004-09-01Bibliographically approved
    4. Complexes of horseradish peroxidase with formate, acetate and carbon monoxide
    Open this publication in new window or tab >>Complexes of horseradish peroxidase with formate, acetate and carbon monoxide
    Show others...
    2005 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 44, no 2, 635-642 p.Article in journal (Refereed) Published
    Abstract [en]

    Carbon monoxide, formate, and acetate interact with horseradish peroxidase (HRP) by binding to subsites within the active site. These ligands also bind to catalases, but their interactions are different in the two types of enzymes. Formate (notionally the “hydrated” form of carbon monoxide) is oxidized to carbon dioxide by compound I in catalase, while no such reaction is reported to occur in HRP, and the CO complex of ferrocatalase can only be obtained indirectly. Here we describe high-resolution crystal structures for HRP in its complexes with carbon monoxide and with formate, and compare these with the previously determined HRP−acetate structure [Berglund, G. I., et al. (2002) Nature 417, 463−468]. A multicrystal X-ray data collection strategy preserved the correct oxidation state of the iron during the experiments. Absorption spectra of the crystals and electron paramagnetic resonance data for the acetate and formate complexes in solution correlate electronic states with the structural results. Formate in ferric HRP and CO in ferrous HRP bind directly to the heme iron with iron−ligand distances of 2.3 and 1.8 Å, respectively. CO does not bind to the ferric iron in the crystal. Acetate bound to ferric HRP stacks parallel with the heme plane with its carboxylate group 3.6 Å from the heme iron, and without an intervening solvent molecule between the iron and acetate. The positions of the oxygen atoms in the bound ligands outline a potential access route for hydrogen peroxide to the iron. We propose that interactions in this channel ensure deprotonation of the proximal oxygen before binding to the heme iron.

    National Category
    Biochemistry and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-91908 (URN)10.1021/bi0483211 (DOI)
    Available from: 2004-09-01 Created: 2004-09-01 Last updated: 2013-12-18Bibliographically approved
  • 19. Chapman, Henry N.
    et al.
    Caleman, Carl
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy, Molecular and condensed matter physics.
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy, Molecular and condensed matter physics. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Diffraction before destruction2014In: Philosophical Transactions of the Royal Society of London. Biological Sciences, ISSN 0962-8436, E-ISSN 1471-2970, Vol. 369, no 1647, 20130313- p.Article in journal (Refereed)
    Abstract [en]

    X-ray free-electron lasers have opened up the possibility of structure determination of protein crystals at room temperature, free of radiation damage. The femtosecond-duration pulses of these sources enable diffraction signals to be collected from samples at doses of 1000 MGy or higher. The sample is vaporized by the intense pulse, but not before the scattering that gives rise to the diffraction pattern takes place. Consequently, only a single flash diffraction pattern can be recorded from a crystal, giving rise to the method of serial crystallography where tens of thousands of patterns are collected from individual crystals that flow across the beam and the patterns are indexed and aggregated into a set of structure factors. The high-dose tolerance and the many-crystal averaging approach allow data to be collected from much smaller crystals than have been examined at synchrotron radiation facilities, even from radiation-sensitive samples. Here, we review the interaction of intense femtosecond X-ray pulses with materials and discuss the implications for structure determination. We identify various dose regimes and conclude that the strongest achievable signals for a given sample are attained at the highest possible dose rates, from highest possible pulse intensities.

  • 20.
    Czapla-Masztafiak, Joanna
    et al.
    Paul Scherrer Inst, Villigen, Switzerland.;Polish Acad Sci, Inst Nucl Phys, Krakow, Poland..
    Szlachetko, Jakub
    Paul Scherrer Inst, Villigen, Switzerland.;Jan Kochanowski Univ Humanities & Sci, Inst Phys, Kielce, Poland..
    Milne, Christopher J.
    Paul Scherrer Inst, Villigen, Switzerland..
    Lipiec, Ewelina
    Sa, Jacinto
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Physical Chemistry. Polish Acad Sci, Inst Phys Chem, Warsaw, Poland..
    Penfold, Thomas J.
    Newcastle Univ, Dept Chem, Bedson Bldg, Newcastle Upon Tyne NE1 7RU, Tyne & Wear, England..
    Huthwelker, Thomas
    Paul Scherrer Inst, Villigen, Switzerland..
    Borca, Camelia
    Paul Scherrer Inst, Villigen, Switzerland..
    Abela, Rafael
    Paul Scherrer Inst, Villigen, Switzerland..
    Kwiatek, Wojciech M.
    Polish Acad Sci, Inst Nucl Phys, Krakow, Poland..
    Investigating DNA Radiation Damage Using X-Ray Absorption Spectroscopy2016In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, no 6, 1304-1311 p.Article in journal (Refereed)
    Abstract [en]

    The biological influence of radiation on living matter has been studied for years; however, several questions about the detailed mechanism of radiation damage formation remain largely unanswered. Among all biomolecules exposed to radiation, DNA plays an important role because any damage to its molecular structure can affect the whole cell and may lead to chromosomal rearrangements resulting in genomic instability or cell death. To identify and characterize damage induced in the DNA sugar-phosphate backbone, in this work we performed x-ray absorption spectroscopy at the P K-edge on DNA irradiated with either UVA light or protons. By combining the experimental results with theoretical calculations, we were able to establish the types and relative ratio of lesions produced by both UVA and protons around the phosphorus atoms in DNA.

  • 21. Danielsson, Ravi
    et al.
    Albertsson, Per-Åke
    Mamedov, Fikret
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström.
    Styring, Stenbjörn
    Quantification of photosystem I and II in different parts of the thylakoid membrane2004In: Biochimica et Biophysica Acta - Bioenergetics, ISSN 0005-2728, E-ISSN 1879-2650, Vol. 1608, 53-61 p.Article in journal (Refereed)
  • 22. D'Arcy, Allan
    et al.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Cowan-Jacob, Sandra W.
    Marsh, May
    Microseed matrix screening for optimization in protein crystallization: what have we learned?2014In: Acta Crystallographica. Section F: Structural Biology and Crystallization Communications, ISSN 1744-3091, E-ISSN 1744-3091, Vol. 70, 1117-1126 p.Article in journal (Refereed)
    Abstract [en]

    Protein crystals obtained in initial screens typically require optimization before they are of X-ray diffraction quality. Seeding is one such optimization method. In classical seeding experiments, the seed crystals are put into new, albeit similar, conditions. The past decade has seen the emergence of an alternative seeding strategy: microseed matrix screening (MMS). In this strategy, the seed crystals are transferred into conditions unrelated to the seed source. Examples of MMS applications from in-house projects and the literature include the generation of multiple crystal forms and different space groups, better diffracting crystals and crystallization of previously uncrystallizable targets. MMS can be implemented robotically, making it a viable option for drug-discovery programs. In conclusion, MMS is a simple, time-and cost-efficient optimization method that is applicable to many recalcitrant crystallization problems.

  • 23.
    Daurer, Benedikt J.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Algorithms for Coherent Diffractive Imaging with X-ray Lasers2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Coherent diffractive imaging (CDI) has become a very popular technique over the past two decades. CDI is a "lensless" imaging method which replaces the objective lens of a conventional microscope by a computational image reconstruction procedure. Its increase in popularity came together with the development of X-ray free-electron lasers (XFELs) which produce extremely bright and coherent X-rays. By facilitating these unique properties, CDI enables structure determination of non-crystalline samples at nanometre resolution and has many applications in structural biology, material science and X-ray optics among others. This work focuses on two specific CDI techniques, flash X-ray diffractive imaging (FXI) on biological samples and X-ray ptychography.

    While the first FXI demonstrations using soft X-rays have been quite promising, they also revealed remaining technical challenges. FXI becomes even more demanding when approaching shorter wavelengths to allow subnanometre resolution imaging. We described one of the first FXI experiments using hard X-rays and characterized the most critical components of such an experiment, namely the properties of X-ray focus, sample delivery and detectors. Based on our findings, we discussed experimental and computational strategies for FXI to overcome its current difficulties and reach its full potential. We deposited the data in the Coherent X-ray Database (CXIDB) and made our data analysis code available in a public repository. We developed algorithms targeted towards the needs of FXI experiments and implemented a software package which enables the analysis of diffraction data in real time.

    X-ray ptychography has developed into a very useful tool for quantitative imaging of complex materials and has found applications in many areas. However, it involves a computational reconstruction step which can be slow. Therefore, we developed a fast GPU-based ptychographic solver and combined it with a framework for real-time data processing which already starts the ptychographic reconstruction process while data is still being collected. This provides immediate feedback to the user and allows high-throughput ptychographic imaging.

    Finally, we have used ptychographic imaging as a method to study the wavefront of a focused XFEL beam under typical FXI conditions. 

    We are convinced that this work on developing strategies and algorithms for FXI and ptychography is a valuable contribution to the development of coherent diffractive imaging. 

    List of papers
    1. Experimental strategies for imaging bioparticles with femtosecond hard X-ray pulses
    Open this publication in new window or tab >>Experimental strategies for imaging bioparticles with femtosecond hard X-ray pulses
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    2017 (English)In: IUCrJ, ISSN 0972-6918, E-ISSN 2052-2525, Vol. 4, 251-262 p.Article in journal (Refereed) Published
    National Category
    Biophysics
    Identifiers
    urn:nbn:se:uu:diva-328529 (URN)10.1107/S2052252517003591 (DOI)
    Projects
    eSSENCE
    Available from: 2017-04-07 Created: 2017-08-25 Last updated: 2017-10-24Bibliographically approved
    2. Hummingbird: monitoring and analyzing flash X-ray imaging experiments in real time
    Open this publication in new window or tab >>Hummingbird: monitoring and analyzing flash X-ray imaging experiments in real time
    2016 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1042-1047 p.Article in journal (Refereed) Published
    National Category
    Biophysics Software Engineering
    Identifiers
    urn:nbn:se:uu:diva-287197 (URN)10.1107/S1600576716005926 (DOI)000377020600036 ()
    Projects
    eSSENCE
    Available from: 2016-04-18 Created: 2016-04-22 Last updated: 2017-10-24Bibliographically approved
    3. SHARP: a distributed GPU-based ptychographic solver
    Open this publication in new window or tab >>SHARP: a distributed GPU-based ptychographic solver
    Show others...
    2016 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1245-1252 p.Article in journal (Refereed) Published
    Abstract [en]

    Ever brighter light sources, fast parallel detectors and advances in phase retrieval methods have made ptychography a practical and popular imaging technique. Compared to previous techniques, ptychography provides superior robustness and resolution at the expense of more advanced and time-consuming data analysis. By taking advantage of massively parallel architectures, high-throughput processing can expedite this analysis and provide microscopists with immediate feedback. These advances allow real-time imaging at wavelength-limited resolution, coupled with a large field of view. This article describes a set of algorithmic and computational methodologies used at the Advanced Light Source and US Department of Energy light sources. These are packaged as a CUDA-based software environment named SHARP (http://camera.lbl.gov/sharp), aimed at providing state-of-the-art high-throughput ptychography reconstructions for the coming era of diffraction-limited light sources.

    Keyword
    coherent X-ray diffractive imaging, ptychography, nanoscience, X-ray microscopy, phase-contrast X-ray imaging
    National Category
    Atom and Molecular Physics and Optics
    Identifiers
    urn:nbn:se:uu:diva-300178 (URN)10.1107/S1600576716008074 (DOI)000382755900015 ()
    Funder
    Swedish Research CouncilSwedish Foundation for Strategic Research
    Available from: 2016-08-04 Created: 2016-08-04 Last updated: 2017-10-24
    4. Nanosurveyor: a framework for real-time data processing
    Open this publication in new window or tab >>Nanosurveyor: a framework for real-time data processing
    Show others...
    2017 (English)In: Advanced Structural and Chemical Imaging, ISSN 2198-0926, Vol. 3, no 7Article in journal (Refereed) Published
    Abstract [en]

    Background: The ever improving brightness of accelerator based sources is enabling novel observations and discov-eries with faster frame rates, larger fields of view, higher resolution, and higher dimensionality.

    Results: Here we present an integrated software/algorithmic framework designed to capitalize on high-throughput experiments through efficient kernels, load-balanced workflows, which are scalable in design. We describe the streamlined processing pipeline of ptychography data analysis.

    Conclusions: The pipeline provides throughput, compression, and resolution as well as rapid feedback to the micro-scope operators

    Keyword
    Streaming, Ptychography
    National Category
    Computer Science
    Identifiers
    urn:nbn:se:uu:diva-317013 (URN)10.1186/s40679-017-0039-0 (DOI)000411110400001 ()28261545 (PubMedID)
    Funder
    Swedish Research CouncilSwedish Foundation for Strategic Research
    Available from: 2017-03-08 Created: 2017-03-08 Last updated: 2017-12-08Bibliographically approved
    5. Wavefront sensing of individual XFEL pulses using ptychography
    Open this publication in new window or tab >>Wavefront sensing of individual XFEL pulses using ptychography
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    The characterization of the wavefront dynamics is important for many X-ray free-electron laser (XFEL) experiments, in particular for coherent diffractive imaging (CDI), as the reconstructed image is always the product of the incoming wavefront with the object. An accurate understanding of the wavefront is also important for any experiment wishing to achieve peak power densities, making use of the tightest possible focal spots. With the use of ptychography we demonstrate high-resolution imaging of the Linac Coherent Light Source (LCLS) beam focused at the endstation for Atomic, Molecular and Optical (AMO) experiments, including its phase and intensity at every plane along its propagation axis, for each individual pulse. Using a mixed-state approach, we have reconstructed the most dominant beam components that constitute an ensemble of pulses, and from the reconstructed components determined their respective contribution in each of the individual pulses. This enabled us to obtain complete wavefront information about each individual pulse. We hope that our findings aid interpretation of data from past and future LCLS experiments and we propose this method to be used routinely for XFEL beam diagnostics. 

    National Category
    Atom and Molecular Physics and Optics
    Identifiers
    urn:nbn:se:uu:diva-329011 (URN)
    Available from: 2017-10-24 Created: 2017-10-24 Last updated: 2017-10-24
  • 24.
    Daurer, Benedikt J.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hantke, Max F.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Nettelblad, Carl
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computational Science. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Maia, Filipe R. N. C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hummingbird: monitoring and analyzing flash X-ray imaging experiments in real time2016In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1042-1047 p.Article in journal (Refereed)
  • 25.
    Daurer, Benedikt J.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Okamoto, Kenta
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Bielecki, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Maia, Filipe R. N. C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Mühlig, Kerstin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Seibert, M. Marvin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hantke, Max F.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Nettelblad, Carl
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computational Science. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Benner, W. Henry
    Svenda, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy, Molecular and Condensed Matter Physics.
    Ekeberg, Tomas
    Loh, N. Duane
    Pietrini, Alberto
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Zani, Alessandro
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Rath, Asawari D.
    Westphal, Daniel
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Kirian, Richard A.
    Awel, Salah
    Wiedorn, Max O.
    van der Schot, Gijs
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Carlsson, Gunilla H.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hasse, Dirk
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Sellberg, Jonas A.
    Barty, Anton
    Andreasson, Jakob
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Boutet, Sébastien
    Williams, Garth
    Koglin, Jason
    Andersson, Inger
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Larsson, Daniel S. D.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Experimental strategies for imaging bioparticles with femtosecond hard X-ray pulses2017In: IUCrJ, ISSN 0972-6918, E-ISSN 2052-2525, Vol. 4, 251-262 p.Article in journal (Refereed)
  • 26.
    de Teran, Hugo Gutierrez
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational Biology and Bioinformatics.
    Understanding ligand binding and receptor selectivity through molecular simulations2015In: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 44, S202-S202 p.Article in journal (Other academic)
  • 27.
    Dogan, Jakob
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Jemth, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Only kinetics can prove conformational selection2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 107, no 8, 1997-1998 p.Article in journal (Other academic)
  • 28.
    Duarte, Fernanda
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Amrein, Beat A.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Blaha-Nelson, David
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Kamerlin, Shina Caroline Lynn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Recent advances in QM/MM free energy calculations using reference potentials2015In: Biochimica et Biophysica Acta - General Subjects, ISSN 0304-4165, E-ISSN 1872-8006, Vol. 1850, no 5, 954-965 p.Article, review/survey (Refereed)
    Abstract [en]

    Background: Recent years have seen enormous progress in the development of methods for modeling (bio)molecular systems. This has allowed for the simulation of ever larger and more complex systems. However, as such complexity increases, the requirements needed for these models to be accurate and physically meaningful become more and more difficult to fulfill. The use of simplified models to describe complex biological systems has long been shown to be an effective way to overcome some of the limitations associated with this computational cost in a rational way. Scope of review: Hybrid QM/MM approaches have rapidly become one of the most popular computational tools for studying chemical reactivity in biomolecular systems. However, the high cost involved in performing high-level QM calculations has limited the applicability of these approaches when calculating free energies of chemical processes. In this review, we present some of the advances in using reference potentials and mean field approximations to accelerate high-level QM/MM calculations. We present illustrative applications of these approaches and discuss challenges and future perspectives for the field. Major conclusions: The use of physically-based simplifications has shown to effectively reduce the cost of high-level QM/MM calculations. In particular, lower-level reference potentials enable one to reduce the cost of expensive free energy calculations, thus expanding the scope of problems that can be addressed. General significance: As was already demonstrated 40 years ago, the usage of simplified models still allows one to obtain cutting edge results with substantially reduced computational cost. This article is part of a Special Issue entitled Recent developments of molecular dynamics.

  • 29.
    Duarte, Fernanda
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Barrozo, Alexandre
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Kamerlin, S. C. Lynn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    Understanding phosphoryl/sulfuryl transfer reactions: from model systems to enzymes2015In: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 44, S165-S165 p.Article in journal (Other academic)
  • 30.
    Ekeberg, Tomas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Flash Diffractive Imaging in Three Dimensions2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    During the last years we have seen the birth of free-electron lasers, a new type of light source ten billion times brighter than syncrotrons and able to produce pulses only a few femtoseconds long. One of the main motivations for building these multi-million dollar machines was the prospect of imaging biological samples such as proteins and viruses in 3D without the need for crystallization or staining. This thesis contains some of the first biological results from free-electron lasers.

    These results include 2D images, both of whole cells and of the giant mimivirus and also con- tains a 3D density map of the mimivirus assembled from diffraction patterns from many virus particles. These are important proof-of-concept experiments but they also mark the point where free-electron lasers start to produce biologically relevant results. The most noteworthy of these results is the unexpectedly non-uniform density distribution of the internals of the mimivirus.

    We also present Hawk, the only open-source software toolkit for analysing single particle diffraction data. The Uppsala-developed program suite supports a wide range fo algorithms and takes advantage of Graphics Processing Units which makes it very computationally efficient.

    Last, the problem introduced by structural variability in samples is discussed. This includes a description of the problem and how it can be overcome, and also how it could be turned into an advantage that allows us to image samples in all of their conformational states.

    List of papers
    1. Three-dimensional structure determination with an X-ray laser
    Open this publication in new window or tab >>Three-dimensional structure determination with an X-ray laser
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Three-dimensional structure determination of a non-crystalline virus has been achieved from a set of randomly oriented continuous diffraction patterns captured with an X-ray laser. Intense, ultra-short X-ray pulses intercepted a beam of single mimivirus particles, producing single particle X-ray diffraction patterns that are assembled into a three-dimensional amplitude distribution based on statistical consistency. Phases are directly retrieved from the assembled Fourier distribution to synthesize a three-dimensional image. The resulting electron density reveals a pseudo-icosahedral asymmetric virion structure with a compartmentalized interior, within which the DNA genome occupies only about a fifth of the volume enclosed by the capsid. Additional electron microscopy data indicate the genome has a chromatin-like fiber structure that has not previously been observed in a virus. 

    Keyword
    Mimivirus, flash diffraction, three dimensional, imaging, CXI
    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:uu:diva-179597 (URN)
    Funder
    EU, European Research Council
    Available from: 2012-08-20 Created: 2012-08-20 Last updated: 2014-09-26
    2. Single mimivirus particles intercepted and imaged with an X-ray laser
    Open this publication in new window or tab >>Single mimivirus particles intercepted and imaged with an X-ray laser
    Show others...
    2011 (English)In: Nature, ISSN 0028-0836, E-ISSN 1476-4687, Vol. 470, no 7332, 78-81 p.Article in journal (Refereed) Published
    Abstract [en]

    X-ray lasers offer new capabilities in understanding the structure of biological systems, complex materials and matter under extreme conditions(1-4). Very short and extremely bright, coherent X-ray pulses can be used to outrun key damage processes and obtain a single diffraction pattern from a large macromolecule, a virus or a cell before the sample explodes and turns into plasma(1). The continuous diffraction pattern of non-crystalline objects permits oversampling and direct phase retrieval(2). Here we show that high-quality diffraction data can be obtained with a single X-ray pulse from a noncrystalline biological sample, a single mimivirus particle, which was injected into the pulsed beam of a hard-X-ray free-electron laser, the Linac Coherent Light Source(5). Calculations indicate that the energy deposited into the virus by the pulse heated the particle to over 100,000 K after the pulse had left the sample. The reconstructed exit wavefront (image) yielded 32-nm full-period resolution in a single exposure and showed no measurable damage. The reconstruction indicates inhomogeneous arrangement of dense material inside the virion. We expect that significantly higher resolutions will be achieved in such experiments with shorter and brighter photon pulses focused to a smaller area. The resolution in such experiments can be further extended for samples available in multiple identical copies.

    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-146069 (URN)10.1038/nature09748 (DOI)000286886400037 ()21293374 (PubMedID)
    Available from: 2011-02-15 Created: 2011-02-15 Last updated: 2017-12-11
    3. Data requirements for single-particle diffractive imaging
    Open this publication in new window or tab >>Data requirements for single-particle diffractive imaging
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Single-shot diffractive imaging with ultra-short and very intense coherent X-ray pulses has become a routine experimental technique at new free-electron-laser facilities. Extension to three-dimensional imaging requires many diffraction pat- terns from identical objects captured in different orientations. These can then be combined into a full three-dimensional Fourier transform of the object. The ori- entation of the particle intercepted by the pulsed X-ray beam is usually unknown. This makes it hard to predict the number of patterns required to fully cover the Fourier space. In this paper we provide formulae to estimate the number of expo- sures required to achieve a given coverage of Fourier space as a function of parti- cle size, resolution and shot noise. 

    National Category
    Natural Sciences
    Identifiers
    urn:nbn:se:uu:diva-179594 (URN)
    Funder
    EU, European Research Council
    Available from: 2012-08-20 Created: 2012-08-20 Last updated: 2014-09-26
    4. Structural variability and the incoherent addition of scattered intensities in single-particle diffraction
    Open this publication in new window or tab >>Structural variability and the incoherent addition of scattered intensities in single-particle diffraction
    Show others...
    2009 (English)In: Physical Review E. Statistical, Nonlinear, and Soft Matter Physics: Statistical Physics, Plasmas, Fluids, and Related Interdisciplinary Topics, ISSN 1063-651X, E-ISSN 1095-3787, Vol. 80, no 3, 031905- p.Article in journal (Refereed) Published
    Abstract [en]

    X-ray lasers may allow structural studies on single particles and biomolecules without crystalline periodicity in the samples. We examine here the effect of sample dynamics as a source of structural heterogeneity on the resolution of the reconstructed image of a small protein molecule. Structures from molecular-dynamics simulations of lysozyme were sampled and aligned. These structures were then used to calculate diffraction patterns corresponding to different dynamic states. The patterns were incoherently summed and the resulting data set was phased using the oversampling method. Reconstructed images of hydrated and dehydrated lysozyme gave resolutions of 3.7 angstrom and 7.6 angstrom, respectively. These are significantly worse than the root-mean-square deviation of the hydrated (2.7 angstrom for all atoms and 1.45 angstrom for C-alpha positions) or dehydrated (3.7 angstrom for all atoms and 2.5 angstrom for C-alpha positions) structures. The noise introduced by structural dynamics and incoherent addition of dissimilar structures restricts the maximum resolution to be expected from direct image reconstruction of dynamic systems. A way of potentially reducing this effect is by grouping dynamic structures into distinct structural substates and solving them separately.

    Keyword
    x-ray-diffraction, electron cascades, proteins, crystallography, resolution, pulses
    National Category
    Physical Sciences
    Identifiers
    urn:nbn:se:uu:diva-111993 (URN)10.1103/PhysRevE.80.031905 (DOI)000270383400104 ()1539-3755 (ISBN)
    Note

    Part 1 501LM Times Cited:0 Cited References Count:32

    Available from: 2010-01-05 Created: 2010-01-05 Last updated: 2017-12-12Bibliographically approved
    5. Hawk: the image reconstruction package for coherent X-ray diffractive imaging
    Open this publication in new window or tab >>Hawk: the image reconstruction package for coherent X-ray diffractive imaging
    2010 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 43, no 6, 1535-1539 p.Article in journal (Other academic) Published
    Abstract [en]

    The past few years have seen a tremendous growth in the field of coherent X-ray diffractive imaging, in large part due to X-ray free-electron lasers which provide a peak brilliance billions of times higher than that of synchrotrons. However, this rapid development in terms of hardware has not been matched on the software side. The release of Hawk is intended to close this gap. To the authors knowledge Hawk is the first publicly available and fully open source software program for reconstructing images from continuous diffraction patterns. The software handles all steps leading from a raw diffraction pattern to a reconstructed two-dimensional image including geometry determination, background correction, masking and phasing. It also includes preliminary three-dimensional support and support for graphics processing units using the Compute Unified Device Architecture, which speeds up processing by orders of magnitude compared to a single central processing unit. Hawk implements numerous algorithms and is easily extended. This, in combination with its open-source licence, provides a platform for other groups to test, develop and distribute their own algorithms.

    Keyword
    computer programs, diffractive imaging, free-electron lasers, Hawk, open-source software, phasing, reconstruction
    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-121927 (URN)10.1107/S0021889810036083 (DOI)000284550900033 ()
    Available from: 2010-03-31 Created: 2010-03-31 Last updated: 2017-12-12Bibliographically approved
    6. Femtosecond diffractive imaging of biological cells
    Open this publication in new window or tab >>Femtosecond diffractive imaging of biological cells
    Show others...
    2010 (English)In: Journal of Physics B: Atomic, Molecular and Optical Physics, ISSN 0953-4075, E-ISSN 1361-6455, Vol. 43, no 19, 194015- p.Article in journal (Refereed) Published
    Abstract [en]

    In a flash diffraction experiment, a short and extremely intense x-ray pulse illuminates the sample to obtain a diffraction pattern before the onset of significant radiation damage. The over-sampled diffraction pattern permits phase retrieval by iterative phasing methods. Flash diffractive imaging was first demonstrated on an inorganic test object (Chapman et al 2006 Nat. Phys. 2 839-43). We report here experiments on biological systems where individual cells were imaged, using single, 10-15 fs soft x-ray pulses at 13.5 nm wavelength from the FLASH free-electron laser in Hamburg. Simulations show that the pulse heated the sample to about 160 000 K but not before an interpretable diffraction pattern could be obtained. The reconstructed projection images return the structures of the intact cells. The simulations suggest that the average displacement of ions and atoms in the hottest surface layers remained below 3 angstrom during the pulse.

    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-147259 (URN)10.1088/0953-4075/43/19/194015 (DOI)000281958100016 ()
    Available from: 2011-02-25 Created: 2011-02-24 Last updated: 2017-12-11Bibliographically approved
  • 31.
    Ekeberg, Tomas
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Engblom, Stefan
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computational Science.
    Liu, Jing
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computational Science.
    Machine learning for ultrafast X-ray diffraction patterns on large-scale GPU clusters2015In: The international journal of high performance computing applications, ISSN 1094-3420, E-ISSN 1741-2846, Vol. 29, 233-243 p.Article in journal (Refereed)
  • 32.
    Ekeberg, Tomas
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Svenda, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Abergel, Chantal
    Maia, Filipe R. N. C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Seltzer, Virginie
    Claverie, Jean-Michel
    Hantke, Max
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Jönsson, Olof
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy, Molecular and Condensed Matter Physics.
    Nettelblad, Carl
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computational Science.
    van der Schot, Gijs
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Liang, Mengning
    DePonte, Daniel P.
    Barty, Anton
    Seibert, M. Marvin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Iwan, Bianca
    Andersson, Inger
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Loh, N. Duane
    Martin, Andrew V.
    Chapman, Henry
    Bostedt, Christoph
    Bozek, John D.
    Ferguson, Ken R.
    Krzywinski, Jacek
    Epp, Sascha W.
    Rolles, Daniel
    Rudenko, Artem
    Hartmann, Robert
    Kimmel, Nils
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Three-dimensional reconstruction of the giant mimivirus particle with an X-ray free-electron laser2015In: Physical Review Letters, ISSN 0031-9007, E-ISSN 1079-7114, Vol. 114, no 9, 098102:1-6 p., 098102Article in journal (Refereed)
  • 33.
    Elf, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Direct Measurements of Transcription Factor Binding and Dissociation at Individual Chromosomal Operators2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 444A-444A p.Article in journal (Other academic)
    Abstract [en]

    I will discuss some of our recent progress in studying transcription factor kinetics at the level of individual molecules in E. coli. I will in particular describe an assay for measuring the rate of dissociation for a LacI repressor from an individual chromosomal operator site. When combined with the corresponding association rate measurement, the assay allows us to test the commonly used assumption that TF kinetics can be considered to be at equilibrium and that the gene expression is proportional to the time the operator is free .

  • 34.
    Elihn, Kristina
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Inorganic Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Structural Chemistry. strukturkemi.
    Otten, F
    Boman, Mats
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Inorganic Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Structural Chemistry. oorganisk kemi.
    Heszler, Peter
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Inorganic Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Structural Chemistry.
    Kruis, FE
    Fissan, H
    Carlsson, Jan-Otto
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Inorganic Chemistry. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Chemistry, Department of Materials Chemistry, Structural Chemistry. oorganisk kemi.
    Size distributions and synthesis of nanoparticles by photolytic dissociation of ferrocene2001In: APPLIED PHYSICS A-MATERIALS SCIENCE & PROCESSING, ISSN 0947-8396, Vol. 72, no 1, 29-34 p.Article in journal (Refereed)
    Abstract [en]

    Iran-containing nanoparticles were made by laser-assisted (ArF excimer laser, lambda = 193 nm) photolytic dissociation of ferrocene (Fe(C5H5)(2) or FeCp2) in argon and an oxygen/argon gas mixture. The particle-size distributions were obtained on-line by u

  • 35.
    Ernst, Aurélie
    et al.
    Karolinska Institut.
    Alkass, Kanar
    Karolinska Institut.
    Bernard, Samuel
    University of Lyon, Villeurbanne, France.
    Salehpour, Mehran
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy.
    Perl, Shira
    NHLBI, NIH, Bethesda, Maryland, USA.
    Tisdale, John
    NHLBI, NIH, Bethesda, Maryland, USA.
    Possnert, Göran
    Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy.
    Druid, Henrik
    Karolinska Institut.
    Frisén, Jonas
    Karolinska Institut.
    Neurogenesis in the Striatum of the Adult Human Brain2014In: Cell, ISSN 0092-8674, E-ISSN 1097-4172, Vol. 156, no 5, 1072-1083 p.Article in journal (Refereed)
    Abstract [en]

    Neurons are added throughout life in the hippocampus and olfactory bulb in most mammals, although humans represent an exception without detectable olfactory bulb neurogenesis. Nevertheless, neuroblasts are generated in the lateral ventricle wall in humans, the neurogenic niche for olfactory bulb neurons in other mammals. We show that, in humans, new neurons integrate adjacent to this neurogenic niche, in the striatum. The neuronal turnover in the striatum appears restricted to interneurons and we show that postnatally generated striatal neurons are preferentially depleted in Huntington’s disease. This demonstrates a unique pattern of neurogenesis in the adult human brain.  

  • 36.
    Flores, Samuel C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Elucidating Ribosomal Translocation with Internal Coordinate Flexible Fitting2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 492A-493A p.Article in journal (Other academic)
    Abstract [en]

    Determining conformational changes of large macromolecules is challenging experimentally and computationally. The ribosome has been observed crystallographically in several states but many others have been seen only by low-resolution methods including cryo-electron microscopy. Meanwhile the crucial dynamics between states remain out of reach of experimental structure determination methods. Most existing computational approaches model complexes at all-atom resolution, at very high cost, or use approximations which lose some of the most interesting dynamical details. I have developed Internal Coordinate Flexible Fitting (ICFF), a multiscale method that uses full atomic forces and flexibility only in key regions of a model, capturing extensive conformational rearrangements at low cost. I use ICFF to turn low-resolution density maps, crystallographic structures, and biochemical information into the largest-scale all-atoms trajectory of ribosomal translocation modeled to date. ICFF is three orders of magnitude faster than the most comparable existing method. The results suggest an intriguing possible mechanism of translocation.

  • 37. Fornander, Louise Helena
    et al.
    Persson, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Fritzsche, Joachim
    Araya, Joshua
    Nevin, Philip
    Beuning, Penny
    Modesti, Mauro
    Frykholm, Karolin
    Westerlund, Fredrik
    Using Nanofluidic Channels to Probe the Dynamics of Rad51-DNA Filaments2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 692A-693A p.Article in journal (Other academic)
    Abstract [en]

    Rad51 is a key protein involved in the strand exchange reaction, a reaction where genetic material is transferred between two homologous DNA strands. Strand exchange is initiated by Rad51 forming a helical filament around single-stranded DNA (ssDNA), and the strand exchange is thereafter executed with a homologous double-stranded DNA (dsDNA). The structure of Rad51-DNA filaments, and also the activity of the strand exchange reaction, is dependent on the presence of ATP and dications, where Ca2+ has been shown to promote a higher degree of strand exchange than Mg2+.

  • 38.
    Fromell, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Yang, Yi
    Gothenburg Univ, Dept Chem & Mol Biol, S-40530 Gothenburg, Sweden..
    Nilsson Ekdahl, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Linnaeus Univ, Linnaeus Ctr Biomat Chem, S-39182 Kalmar, Sweden..
    Nilsson, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Linnaeus Univ, Linnaeus Ctr Biomat Chem, S-39182 Kalmar, Sweden..
    Berglin, Mattias
    Gothenburg Univ, Dept Chem & Mol Biol, S-40530 Gothenburg, Sweden.;RISE Res Inst Sweden Chem Mat & Surfaces, S-50115 Boras, Sweden..
    Elwing, Hans
    Gothenburg Univ, Dept Chem & Mol Biol, S-40530 Gothenburg, Sweden..
    Absence of conformational change in complement factor 3 and factor XII adsorbed to acrylate polymers is related to a high degree of polymer backbone flexibility2017In: Biointerphases, ISSN 1934-8630, E-ISSN 1559-4106, Vol. 12, no 2, 02D417Article in journal (Refereed)
    Abstract [en]

    In previous investigations, the authors have examined the adsorption of albumin, immunoglobulin, and fibrinogen to a series of acrylate polymers with different backbone and side-group flexibility. The authors showed that protein adsorption to acrylates with high flexibility, such as poly(lauryl methacrylate) (PLMA), tends to preserve native conformation. In the present study, the authors have continued this work by examining the conformational changes that occur during the binding of complement factor 3 (C3) and coagulation factor XII (FXII). Native C3 adsorbed readily to all solid surfaces tested, including a series of acrylate surfaces of varying backbone flexibility. However, a monoclonal antibody recognizing a "hidden" epitope of C3 (only exposed during C3 activation or denaturation) bound to the C3 on the rigid acrylate surfaces or on polystyrene (also rigid), but not to C3 on the flexible PLMA, indicating that varying degrees of conformational change had occurred with binding to different surfaces. Similarly, FXII was activated only on the rigid poly(butyl methacrylate) surface, as assessed by the formation of FXIIa-antithrombin (AT) complexes; in contrast, it remained in its native form on the flexible PLMA surface. The authors also found that water wettability hysteresis, defined as the difference between the advancing and receding contact angles, was highest for the PLMA surface, indicating that a dynamic change in the interface polymer structure may help protect the adsorbed protein from conformational changes and denaturation.

  • 39. Frykholm, Karolin
    et al.
    Alizadehheidari, Mohammadreza
    Fritzsche, Joachim
    Wigenius, Jens
    Nevin, Philip
    Araya, Joshua
    Beuning, Penny
    Modesti, Mauro
    Persson, Fredrik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
    Westerlund, Fredrik
    Probing Physical Properties of a DNA-Protein Complex Using Nanofluidic Channels2014In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 106, no 2, 428A-429A p.Article in journal (Other academic)
    Abstract [en]

    Nanofluidic channels have become an important tool to investigate single DNA molecules both from a fundamental polymer physics perspective as well as in e.g. optical mapping techniques. However, less effort has been made to study DNA-protein complexes. A main reason is that the extreme surface-to-volume ratio in the nanochannels causes most proteins to stick to the channel walls. We have recently overcome this problem by coating the channels with a lipid bilayer, thereby eliminating sticking.

  • 40.
    Galichanin, Konstantin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Löfgren, Stefan
    Söderberg, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Ophthalmology.
    Cataract after repeated daily in vivo exposure to ultraviolet radiation2014In: Health Physics, ISSN 0017-9078, E-ISSN 1538-5159, Vol. 107, no 6, 523-529 p.Article in journal (Refereed)
    Abstract [en]

    Epidemiological data indicate a correlation between lifelong exposure to ultraviolet radiation and cortical cataract. However, there is no quantitative experimental data on the effect of daily repeated in vivo exposures of the eye to UVR. Therefore, this experiment was designed to verify whether the dose additivity for UVR exposures holds through periods of time up to 30 d. Eighty rats were conditioned to a rat restrainer 5 d prior to exposure. All animals were divided into four exposure period groups of 1, 3, 10, and 30 d of exposure to UVR. Each exposure period group of 20 animals was randomly divided into five cumulated UVR dose subgroups. Eighteen-wk-old non-anesthetized albino Sprague-Dawley rats were exposed daily to UVR-300 nm for 15 min. One week after the last exposure, animals were sacrificed. The lenses were extracted for macroscopic imaging of dark-field anatomy, and degree of cataract was quantified by measurement of the intensity of forward lens light scattering. Maximum tolerable dose (MTD2.3:16), a statistically defined standard for sensitivity for the threshold for UVR cataract, was estimated for each exposure period. Exposed lenses developed cataract with varying appearance on the anterior surface. Single low doses of UVR accumulated to cause cataract during periods up to 30 d. MTD2.3:16 for 1, 3, 10, and 30 d of repeated exposures was estimated to 4.70, 4.74, 4.80, and 6.00 kJ m, respectively. In conclusion, the lens sensitivity to UVR-B for 18-wk-old Sprague-Dawley rats decreases with the increasing number of days being exposed. 

  • 41.
    Gandasi, Nikhil R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Barg, Sebastian
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Quantitative Imaging of the Exocytosis Machinery Assembly2015In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, no 2, 102A-102A p.Article in journal (Other academic)
  • 42. Gasanov, Ralphreed
    et al.
    Aliyeva, Samira
    Mamedov, Fikret
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Molecular Biomimetics.
    Delayed fluorescence in a millisecond range: a probe for the donor side-induced photoinhibition in photosystem II2015In: Photosynthesis: Basics to applications / [ed] S. Itoh, P. Mohanty, K.N. Guruprasad, New Delhi, India: I.K. International Publishing House Pvt. Ltd. , 2015, 101-107 p.Chapter in book (Refereed)
  • 43. Gerlach, Samantha L.
    et al.
    Göransson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Kaas, Quentin
    Craik, David J.
    Mondal, Debasis
    Gruber, Christian W.
    A Systematic Approach to Document Cyclotide Distribution in Plant Species from Genomic, Transcriptomic, and Peptidomic Analysis2013In: Biopolymers, ISSN 0006-3525, E-ISSN 1097-0282, Vol. 100, no 5, 433-437 p.Article in journal (Refereed)
    Abstract [en]

    Cyclotides are a large family of plant peptides characterized by their cyclic cystine knot composed of a circular backbone and three disulfide bonds that impart exceptional stability. They, and several acyclic variants, have been isolated from plants within the Rubiaceae, Violaceae, Cucurbitaceae, Fabaceae, Solanaceae, and Poaceae families. A variety of chemical and genetic approaches have been applied for the discovery and characterization of cyclotides. As investigations of cyclotide expression, distribution, and phylogeny rapidly increase, the authors have proposed the inclusion of information pertaining to plant species that have been analyzed but do not appear to express cyclotides into the CyBase database. CyBase is dedicated to providing web tools and information about cyclic peptides and proteins to the scientific community. Including detailed information about sampling and analysis parameters of plant species that have been investigated but not published elsewhere should assist in the process of selecting species for establishing new cyclotide discovery projects, as well as for detailed reanalysis using alternative technical approaches. In summary, the collection and deposition of all plant species that have been examined (whether cyclotides have been found or not) would help to impart a deeper understanding of cyclotide discovery, evolution, and physiological function.

  • 44. Gerlach, Samantha L.
    et al.
    Yeshak, Mariamawit
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Göransson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Roy, Upal
    Izadpanah, Reza
    Mondal, Debasis
    Cycloviolacin O2 (CyO2) Suppresses Productive Infection and Augments the Antiviral Efficacy of Nelfinavir in HIV-1 Infected Monocytic Cells2013In: Biopolymers, ISSN 0006-3525, E-ISSN 1097-0282, Vol. 100, no 5, 471-479 p.Article in journal (Refereed)
    Abstract [en]

    Human immunodeficiency virus type-1 (HIV-1), the etiologic agent of acquired immune deficiency syndrome (AIDS), is a global pandemic causing millions of deaths annually. Highly active antiretroviral therapy (HAART) greatly enhances lifespan but eventually causes debilitating side effects, in part, due to their chronic administration required to suppress HIV-1 replication. If treatment is discontinued, viral suppression is lost and dormant replication-competent monocytic cell reservoirs become reactivated, leading to viral recrudescence and progression to AIDS. Therefore, novel strategies to circumvent obstacles to HIV-1 therapy are critically needed. We evaluated the potentially therapeutic effects of cycloviolacin O2 (CyO2) on cell viability (MTTassay), membrane disruption (SYTOX Green uptake), p24 production [enzyme-linked immunosorbent assays (ELISA)], and proviral integration (PCR amplification) in U1 cells; a monocytic cell model of HIV-1 latency and reactivation. We demonstrate, for the first time, that CyO2 (0.5-5.0 mu M) kills productively infected cells. Sub-toxic concentrations (< 0.5 mu M) of CyO2 disrupted plasma membranes in both latently-infected and productively-infected U1 cells and enhanced the antiviral efficacy of nelfinavir, a HIV-1 protease inhibitor (HPI). Interestingly, CyO2 also decreased virus production by activated U1 cells; however, this effect was not due to suppression of integrated provirus in U1 cells. This suggested that, in addition to the known pore-forming ability of cyclotides, a novel mode of antiviral activity may exist for CyO2. Our data indicate that CyO2 may be a promising candidate for the targeting HIV-1 reservoirs in monocytes, and their inclusion in adjuvant therapy approaches may augment the efficacy of HPIs and ultimately facilitate virus elimination.

  • 45. Gianni, Stefano
    et al.
    Dogan, Jakob
    Jemth, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Distinguishing induced fit from conformational selection2014In: Biophysical Chemistry, ISSN 0301-4622, E-ISSN 1873-4200, Vol. 189, 33-39 p.Article in journal (Refereed)
    Abstract [en]

    The interactions between proteins and ligands often involve a conformational change in the protein. This conformational change can occur before (conformational selection) or after (induced fit) the association with ligand. It is often very difficult to distinguish induced fit from conformational selection when hyperbolic binding kinetics are observed. In light of a recent paper in this journal (Vogt et al., Biophys. Chem., 186, 2014, 13-21) and the current interest in binding mechanisms emerging from observed sampling of distinct conformations in protein domains, as well as from the field of intrinsically disordered proteins, we here describe a kinetic method that, at least in some cases, unequivocally distinguishes induced fit from conformational selection. The method relies on measuring the observed rate constant A. for binding and varying both the protein and the ligand in separate experiments. Whereas induced fit always yields a hyperbolic dependence of increasing A. values, the conformational selection mechanism gives rise to distinct kinetics when the ligand and protein (displaying the conformational change) concentration is varied in separate experiments. We provide examples from the literature and discuss the limitations of the approach.

  • 46.
    Gomez-Llobregat, Jordi
    et al.
    Stockholm Univ, Dept Biochem & Biophys, Ctr Biomembrane Res, S-10691 Stockholm, Sweden..
    Elias-Wolff, Federico
    Stockholm Univ, Dept Biochem & Biophys, Ctr Biomembrane Res, S-10691 Stockholm, Sweden..
    Lindén, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Anisotropic Membrane Curvature Sensing by Amphipathic Peptides2016In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, no 1, 197-204 p.Article in journal (Refereed)
    Abstract [en]

    Many proteins and peptides have an intrinsic capacity to sense and induce membrane curvature, and play crucial roles for organizing and remodeling cell membranes. However, the molecular driving forces behind these processes are not well understood. Here, we describe an approach to study curvature sensing by simulating the interactions of single molecules with a buckled lipid bilayer. We analyze three amphipathic antimicrobial peptides, a class of membrane-associated molecules that specifically target and destabilize bacterial membranes, and find qualitatively different sensing characteristics that would be difficult to resolve with other methods. Our findings provide evidence for direction-dependent curvature sensing mechanisms in amphipathic peptides and challenge existing theories of hydrophobic insertion. The buckling approach is generally applicable to a wide range of curvature-sensing molecules, and our results provide strong motivation to develop new experimental methods to track position and orientation of membrane proteins.

  • 47. Gromova, Arina
    Voltage-gated K+ channel modulation by resin-acid derivatives - a computational study2017Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Voltage-gated K+ (Kv) channels are known to cause serious disease upon their malfunction. Kv channels desensitised to voltage show inability to fully repolarise the membrane in excitable cells, which can make the membrane hyperexcited and in turn cause seizures such as in epilepsy, periodic ataxia or heart arrhythmia. Therefore, enhancers of Kv channels could serve as potential drugs. Some of these enhancers are polyunsaturated fatty acids and resin-acids which bind at the proteinlipid surface and affect the movement of the voltage sensor in the channel by a mechanism called the lipoelectric effect. To explore the lipoelectric modulation mechanism, we have performed an extensive computational study including docking and molecular dynamics simulations on resin-acid derivatives added to a model potassium channel called Shaker. Four derivatives, Wu32 and Wu50 that excite the channel and thus induce repolarisation of the membrane, as well as Wu18 and Wu27, who were found to be non-potent in previous experimental studies, have helped to point out a novel binding site in Shaker. The site is located between the pore and voltage-sensing domain of the channel and is in direct contact with the first gating charge arginine, R1, and the residue W454. We hypothesize that it is possible for resinacid derivatives to directly bind to the voltage-sensor when it is in an activated state, prolonging the time Shaker stays open. Further experimental studies on Shaker and human homologs are now needed to test our hypothesis. Therefore, we suggest recording the sensitivity of Shaker towards potent derivatives in combination with mutations of W454. If our findings of the novel binding site are correct, the suitability of Shaker as a model system for human Kv channel modulation by lipoelectric modulators can be questioned as W454 is replaced by small hydrophobic side chains in mammalian Shaker homologs.

  • 48. Han, Guangye
    et al.
    Morvaridi, Susan
    Ho, Felix
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Molecular Biomimetics.
    Mamedov, Fikret
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Molecular Biomimetics.
    Styring, Stenbjörn
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Molecular Biomimetics.
    S-state dependence of misses in the OEC probed by EPR spectroscopy of individual S-states2008In: Photosynthesis. Energy from the Sun / [ed] Allen J.F., Gant E., Golbeck J.H. and Osmond B., Springer , 2008, 419-422 p.Conference paper (Refereed)
  • 49.
    Hantke, Max F.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Ekeberg, Tomas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Maia, Filipe R. N. C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Condor: a simulation tool for flash X-ray imaging2016In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1356-1362 p.Article, review/survey (Refereed)
    Abstract [en]

    Flash X-ray imaging has the potential to determine structures down to molecular resolution without the need for crystallization. The ability to accurately predict the diffraction signal and to identify the optimal experimental configuration within the limits of the instrument is important for successful data collection. This article introduces Condor, an open-source simulation tool to predict X-ray far-field scattering amplitudes of isolated particles for customized experimental designs and samples, which the user defines by an atomic or a refractive index model. The software enables researchers to test whether their envisaged imaging experiment is feasible, and to optimize critical parameters for reaching the best possible result. It also aims to support researchers who intend to create or advance reconstruction algorithms by simulating realistic test data. Condor is designed to be easy to use and can be either installed as a Python package or used from its web interface (http://lmb.icm.uu.se/condor). X-ray free-electron lasers have high running costs and beam time at these facilities is precious. Data quality can be substantially improved by using simulations to guide the experimental design and simplify data analysis.

  • 50.
    Hantke, Max F.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hasse, Dirk
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Ekeberg, Tomas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    John, Katja
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Svenda, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Loh, Duane
    Martin, Andrew V.
    Timneanu, Nicusor
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics. Uppsala University, Disciplinary Domain of Science and Technology, Physics, Department of Physics and Astronomy, Molecular and Condensed Matter Physics.
    Larsson, Daniel S.D.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    van der Schot, Gijs
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Carlsson, Gunilla H.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Ingelman, Margareta
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Andreasson, Jakob
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Westphal, Daniel
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Iwan, Bianca
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Uetrecht, Charlotte
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Bielecki, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Liang, Mengning
    Stellato, Francesco
    DePonte, Daniel P.
    Bari, Sadia
    Hartmann, Robert
    Kimmel, Nils
    Kirian, Richard A.
    Seibert, M. Marvin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Mühlig, Kerstin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Schorb, Sebastian
    Ferguson, Ken
    Bostedt, Christoph
    Carron, Sebastian
    Bozek, John D.
    Rolles, Daniel
    Rudenko, Artem
    Foucar, Lutz
    Epp, Sascha W.
    Chapman, Henry N.
    Barty, Anton
    Andersson, Inger
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Hajdu, Janos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    Maia, Filipe R.N.C.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.
    A data set from flash X-ray imaging of carboxysomes2016In: Scientific Data, E-ISSN 2052-4463, Vol. 3, 160061Article in journal (Refereed)
    Abstract [en]

    Ultra-intense femtosecond X-ray pulses from X-ray lasers permit structural studies on single particles and biomolecules without crystals. We present a large data set on inherently heterogeneous, polyhedral carboxysome particles. Carboxysomes are cell organelles that vary in size and facilitate up to 40% of Earth’s carbon fixation by cyanobacteria and certain proteobacteria. Variation in size hinders crystallization. Carboxysomes appear icosahedral in the electron microscope. A protein shell encapsulates a large number of Rubisco molecules in paracrystalline arrays inside the organelle. We used carboxysomes with a mean diameter of 115±26 nm from Halothiobacillus neapolitanus. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min. Every diffraction pattern is a unique structure measurement and high-throughput imaging allows sampling the space of structural variability. The different structures can be separated and phased directly from the diffraction data and open a way for accurate, high-throughput studies on structures and structural heterogeneity in biology and elsewhere.

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