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  • 1. Abola, EE
    et al.
    Bairoch, A
    Barker, WC
    Beck, S
    Benson, DA
    Berman, H
    Cantor, C
    Cantor, C
    Doubet, S
    Hubbard, TJP
    Jones, T. A.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, G J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kolaskar, AS
    van Kuik, A
    Lesk, A M
    Mewes, H W
    Neuhaus, D
    Pfeiffer, F
    Ten Eyck, LF
    Simpson, RJ
    Stoesser, G
    Sussman, J L
    Tateno, Y
    Tsugita, A
    Ulrich, EL
    Vliegenthart, JFG
    Quality control in databanks for molecular biology2000In: BioEssays, Vol. 22, p. 1024-1034Article, review/survey (Other (popular scientific, debate etc.))
  • 2.
    Almlöf, Martin
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Aqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Smalås, Arne O
    Brandsdal, Björn O
    Probing the effect of point mutations at protein-protein interfaces with free energy calculations.2006In: Biophys J, ISSN 0006-3495, Vol. 90, no 2, p. 433-42Article in journal (Refereed)
  • 3.
    Almlöf, Martin
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Brandsdal, Bjørn O
    Aqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Binding affinity prediction with different force fields: examination of the2004In: J Comput Chem, ISSN 0192-8651, Vol. 25, no 10, p. 1242-54Article in journal (Refereed)
  • 4.
    Andér, Martin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Luzhkov, Victor B.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Ligand binding to the voltage-gated Kv1.5 potassium channel in the open state - Docking and computer simulations of a homology model2008In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 94, no 3, p. 820-831Article in journal (Refereed)
    Abstract [en]

    The binding of blockers to the human voltage-gated Kv1.5 potassium ion channel is investigated using a three-step procedure consisting of homology modeling, automated docking, and binding free energy calculations from molecular dynamics simulations, in combination with the linear interaction energy method. A reliable homology model of Kv1.5 is constructed using the recently published crystal structure of the Kv1.2 channel as a template. This model is expected to be significantly more accurate than earlier ones based on less similar templates. Using the three-dimensional homology model, a series of blockers with known affinities are docked into the cavity of the ion channel and their free energies of binding are calculated. The predicted binding free energies are in very good agreement with experimental data and the binding is predicted to be mainly achieved through nonpolar interactions, whereas the relatively small differences in the polar contribution determine the specificity. Apart from confirming the importance of residues V505, I508, V512, and V516 for ligand binding in the cavity, the results also show that A509 and P513 contribute significantly to the nonpolar binding interactions. Furthermore, we find that pharmacophore models based only on optimized free ligand conformations may not necessarily capture the geometric features of ligands bound to the channel cavity. The calculations herein give a detailed structural and energetic picture of blocker binding to Kv1.5 and this model should thus be useful for further ligand design efforts.

  • 5.
    Andér, Martin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Does glutamine methylation affect the intrinsic conformation of the universally conserved GGQ motif in ribosomal release factors?2009In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 48, no 15, p. 3483-3489Article in journal (Refereed)
    Abstract [en]

    The GGQ motif is the only universally conserved feature of ribosomal class 1 release factors. Mutational experiments and structural studies have suggested that the glutamine residue of the GGQ motif Q 185 in human eRF1 numbering) is critical for catalysis of the termination   reaction on the ribosome. Furthermore, it has been established that Q185 is NE methylated in prokaryotes as well as eukaryotes, and that methylation significantly enhances the catalytic activity. It is, however, not known whether this methylation affects the intrinsic   structure of the free release factor, which could be important for its interaction with the ribosome. In this work, we report molecular dynamics simulations, starting from 25 different NMR structures of human eRF1, in addressing this problem. The results show that there is   no such structural effect on the free release factor caused by the NE methylation of Q185, suggesting that its role is intimately associated with the ribosome environment.

  • 6. Becker, D
    et al.
    Braet, C
    Brumer, H
    Claeyssens, M
    Divne, C
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Fagerström, VM
    Harris, M
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Jones, TA
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, GJ
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Koivula, A
    Mahdi, S
    Piens, K
    Sinnottt, ML
    Ståhlberg, J
    Teeri, TT
    Underwood, M
    Wohlfart, G
    Engineering of a glycosidase Family 7 cellobiohydrolase to more alkaline pH optimum: the pH behaviour of Trichoderma reesei CeI7A and its E223S/A224H/L225V/T226A/D262G mutant2001In: Biochemical Journal, Vol. 356, p. 19-30Article in journal (Refereed)
    Abstract [en]

    The crystal structures of Family 7 glycohydrolases suggest that a histidine residue near the acid/base catalyst could account for the higher pH optimum of the Humicola insolens endoglucanase Cel7B, than the corresponding Trichoderma reesei enzymes. Modelling studies indicated that introduction of histidine at the homologous position in T. reesei Cel7A (Ala(224)) required additional changes to accommodate the bulkier histidine side chain. X-ray crystallography of the catalytic domain of the E223S/A224H/L225V/T226A/D262G mutant reveals that major differences from the wild-type are confined to the mutations themselves. The introduced histidine residue is in plane with its counterpart in H. insolens Cel7B, but is 1.0 A (=0.1 nm) closer to the acid/base Glu(217) residue, with a 3.1 A contact between N(epsilon2) and O(epsilon1). The pH variation of k(cat)/K(m) for 3,4-dinitrophenyl lactoside hydrolysis was accurately bell-shaped for both wild-type and mutant, with pK(1) shifting from 2.22+/-0.03 in the wild-type to 3.19+/-0.03 in the mutant, and pK(2) shifting from 5.99+/-0.02 to 6.78+/-0.02. With this poor substrate, the ionizations probably represent those of the free enzyme. The relative k(cat) for 2-chloro-4-nitrophenyl lactoside showed similar behaviour. The shift in the mutant pH optimum was associated with lower k(cat)/K(m) values for both lactosides and cellobiosides, and a marginally lower stability. However, k(cat) values for cellobiosides are higher for the mutant. This we attribute to reduced non-productive binding in the +1 and +2 subsites; inhibition by cellobiose is certainly relieved in the mutant. The weaker binding of cellobiose is due to the loss of two water-mediated hydrogen bonds.

  • 7.
    Bergfors, T
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, G J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Jones, T A
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Crystallization and preliminary X-ray analysis of recombinant bovine cellular retinoic acid-binding protein.1994In: Acta Crystallogr D Biol Crystallogr, ISSN 0907-4449, Vol. 50, no Pt 4, p. 370-4Article in journal (Refereed)
    Abstract [en]

    Crystals of bovine cellular retinoic acid-binding protein (CRABPI) have been grown from protein expressed in E. coli. Two different crystal forms were obtained. Crystals containing protein with the ligand all-trans retinoic acid belong to space group P4(1) or P4(3), a = b = 41.36, c = 202.71 A and diffract to 2.5 A. Crystals of CRABP with the synthetic retinoid analogue Am80 diffract to 1.9 A with space group P2(1) and cell dimensions a = 37.03, b = 105.93, c = 40.31 A, beta = 110.28 degrees. Considerations in the crystallization of proteins with light-sensitive ligands are discussed.

  • 8.
    Bergfors, Terese
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. strukturell molekylärbiologi.
    Automated liquid handling systems for submicroliter crystallization2007In: Protein Crystallization Strategies for Structural Genomics, International University Line, La Jolla California , 2007, p. 57-86Chapter in book (Refereed)
  • 9.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Automated liquid-handling systems for submicroliter crystallization2007In: Protein Crystallization Strategies for Structural Genomics / [ed] Naomi E. Chayen, La Jolla, California: International University Line , 2007, p. 57-73Chapter in book (Other academic)
  • 10.
    Bergfors, Terese
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Crystallization strategy at the Uppsala University RAPID Center2006In: Synchrotron Radiation Science & Technology, Vol. 13, p. 5-9Article in journal (Refereed)
  • 11.
    Bergfors, Terese
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Screening and optimization methods for non-automated crystallization laboratories2006In: Methods in Molecular Biology: Macromolecular Crystallography Protocols, Humana Press, New Jersey, USA , 2006, p. 131-151Chapter in book (Refereed)
  • 12.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Succeeding with seeding: some practical advice2007In: Evolving Methods for Macromolecular Crystallography / [ed] Read, Randy J.; Sussman, Joel L., 2007, p. 1-10Conference paper (Refereed)
    Abstract [en]

    Seeding is a powerful and versatile method for optimizing crystal growth conditions. This article discusses, from a practical point of view, what seeding is, the selection and transfer of seeds, and into what conditions they should be transferred. The most common causes of failures in seeding experiments are also analyzed.

  • 13.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    The RAPID Crystallization Strategy for Structure-Based Inhibitor Design2008In: From Molecules to Medicines: Structure of biological macromolecules and its relevance in combating new diseases and bioterrorism, Amsterdam and Dordrecht: IOS Press and Springer , 2008, p. 11-19Chapter in book (Other academic)
  • 14. Bjelic, Sinisa
    et al.
    Aqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Catalysis and linear free energy relationships in aspartic proteases.2006In: Biochemistry, ISSN 0006-2960, Vol. 45, no 25, p. 7709-23Article in journal (Refereed)
  • 15.
    Bjelic, Sinisa
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Aqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Computational prediction of structure, substrate binding mode, mechanism, and rate for a malaria protease with a novel type of active site.2004In: Biochemistry, ISSN 0006-2960, Vol. 43, no 46, p. 14521-8Article in journal (Other scientific)
  • 16.
    Bjelic, Sinisa
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Brandsdal, Bjørn O
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Cold adaptation of enzyme reaction rates2008In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 47, no 38, p. 10049-10057Article in journal (Refereed)
    Abstract [en]

    A major issue for organisms living at extreme temperatures is to preserve both stability and activity of their enzymes. Cold-adapted enzymes generally have a reduced thermal stability, to counteract freezing, and show a lower enthalpy and a more negative entropy of activation compared to mesophilic and thermophilic homologues. Such a balance of thermodynamic activation parameters can make the reaction rate decrease more linearly, rather than exponentially, as the temperature is lowered, but the structural basis for rate optimization toward low working temperatures remains unclear. In order to computationally address this problem, it is clear that reaction simulations rather than standard molecular dynamics calculations are needed. We have thus carried out extensive computer simulations of the keto-enol(ate) isomerization steps in differently adapted citrate synthases to explore the structure-function relationships behind catalytic rate adaptation to different temperatures. The calculations reproduce the absolute rates of the psychrophilic and mesophilic enzymes at 300 K, as well as the lower enthalpy and more negative entropy of activation of the cold-adapted enzyme, where the latter simulation result is obtained from high-precision Arrhenius plots. The overall catalytic effect originates from electrostatic stabilization of the transition state and enolate and the reduction of reorganization free energy. The simulations, however, show psychrophilic, mesophilic, and hyperthermophilic citrate synthases to have increasingly stronger electrostatic stabilization of the transition state, while the energetic penalty in terms of internal protein interactions follows the reverse order with the cold-adapted enzyme having the most favorable energy term. The lower activation enthalpy and more negative activation entropy observed for cold-adapted enzymes are found to be associated with a decreased protein stiffness. The origin of this effect is, however, not localized to the active site but to other regions of the protein structure.

  • 17. Bollati, Michela
    et al.
    Alvarez, Karin
    Assenberg, Rene
    Baronti, Cecile
    Canard, Bruno
    Cook, Shelley
    Coutard, Bruno
    Decroly, Etienne
    de Lamballerie, Xavier
    Gould, Ernest A.
    Grard, Gilda
    Grimes, Jonathan M.
    Hilgenfeld, Rolf
    Jansson, Anna M.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Malet, Helene
    Mancini, Erika J.
    Mastrangelo, Eloise
    Mattevi, Andrea
    Milani, Mario
    Moureau, Gregory
    Neyts, Johan
    Owens, Raymond J.
    Ren, Jingshan
    Selisko, Barbara
    Speroni, Silvia
    Steuber, Holger
    Stuart, David I.
    Unge, Torsten
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Bolognesi, Martino
    Structure and functionality in flavivirus NS-proteins: perspectives for drug design2010In: Antiviral Research, ISSN 0166-3542, E-ISSN 1872-9096, Vol. 87, no 2, p. 125-148Article, review/survey (Refereed)
    Abstract [en]

    Flaviviridae are small enveloped viruses hosting a positive-sense single-stranded RNA genome. Besides yellow fever virus, a landmark case in the history of virology, members of the Flavivirus genus, such as West Nile virus and dengue virus, are increasingly gaining attention due to their re-emergence and incidence in different areas of the world. Additional environmental and demographic considerations suggest that novel or known flaviviruses will continue to emerge in the future. Nevertheless, up to few years ago flaviviruses were considered low interest candidates for drug design. At the start of the European Union VIZIER Project, in 2004, just two crystal structures of protein domains from the flaviviral replication machinery were known. Such pioneering studies, however, indicated the flaviviral replication complex as a promising target for the development of antiviral compounds. Here we review structural and functional aspects emerging from the characterization of two main components (NS3 and NS5 proteins) of the flavivirus replication complex. Most of the reviewed results were achieved within the European Union VIZIER Project, and cover topics that span from viral genomics to structural biology and inhibition mechanisms. The ultimate aim of the reported approaches is to shed light on the design and development of antiviral drug leads.

  • 18.
    Brandsdal, B., Österberg, F., Almlöf, M., Feierberg, I., Luzhkov, V.B.
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Free Energy Calculations and Ligand Binding2003In: Adv. Prot. Chem., Vol. 66, p. 123-Article in journal (Refereed)
  • 19. Brandsdal, Björn O
    et al.
    Smalås, Arne O
    Åqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Free energy calculations show that acidic P1 variants undergo large pKa shifts upon binding to trypsin.2006In: Proteins, ISSN 1097-0134, Vol. 64, no 3, p. 740-8Article in journal (Refereed)
  • 20.
    Brandsdal, B.O., Smalås, A.O
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Electrostatic Effects Play a Central Role in Cold Adaptation of Trypsin2001In: FEBS Lett., Vol. 499, p. 171-Article in journal (Refereed)
  • 21.
    Brandsdal, B.O.
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Smalås, A.O.
    Computational Analysis of Binding of P1-Variants to Trypsin2001In: Protein Sci., Vol. 10, p. 1584-Article in journal (Refereed)
  • 22. Carlsson, Jens
    et al.
    Andér, Martin
    Nervall, Martin
    Aqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Continuum solvation models in the linear interaction energy method.2006In: J Phys Chem B Condens Matter Mater Surf Interfaces Biophys, ISSN 1520-6106, Vol. 110, no 24, p. 12034-41Article in journal (Refereed)
  • 23.
    Carlsson, Jens
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Boukharta, Lars
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Combining docking, molecular dynamics and the linear interaction energy method to predict binding modes and affinities for non-nucleoside inhibitors to HIV-1 reverse transcriptase2008In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 51, no 9, p. 2648-56Article in journal (Refereed)
    Abstract [en]

    Docking, scoring, molecular dynamics (MD), and the linear interaction energy (LIE) method are used here to predict binding modes and affinities for a set of 43 non-nucleoside inhibitors to HIV-1 reverse transcriptase. Starting from a crystallographic structure, the binding modes of 43 inhibitors are predicted using automated docking. The Goldscore scoring function and the LIE method are then used to determine the relative binding free energies for the inhibitors. The Goldscore scoring function does not reproduce the relative binding affinities for the inhibitors, while the standard parametrization of the LIE method reproduces the experimental binding free energies for 39 inhibitors with an R (2) = 0.70 and an unsigned average error of 0.8 kcal/mol. The present calculations provide a validation of the combination of docking, MD, and LIE as a powerful tool in structure-based drug design, and the methodology is easily scalable for attaining a higher throughput of compounds.

  • 24. Carlsson, Jens
    et al.
    Åqvist, Johan
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Absolute and Relative Entropies from Computer Simulation with Applications to Ligand Binding2005In: J. Phys. Chem. B, Vol. 109, p. 6448-Article in journal (Refereed)
  • 25.
    Carlsson, Jens
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Calculations of solute and solvent entropies from molecular dynamics simulations2006In: Physical Chemistry, Chemical Physics - PCCP, ISSN 1463-9076, E-ISSN 1463-9084, Vol. 8, no 46, p. 5385-5395Article in journal (Refereed)
    Abstract [en]

    The translational, rotational and conformational ( vibrational) entropy contributions to ligand-receptor binding free energies are analyzed within the standard formulation of statistical thermodynamics. It is shown that the partitioning of the binding entropy into different components is to some extent arbitrary, but an appropriate method to calculate both translational and rotational entropy contributions to noncovalent association is by estimating the configurational volumes of the ligand in the bound and free states. Different approaches to calculating solute entropies using free energy perturbation calculations, configurational volumes based on root-mean-square fluctuations and covariance matrix based quasiharmonic analysis are illustrated for some simple molecular systems. Numerical examples for the different contributions demonstrate that theoretically derived results are well reproduced by the approximations. Calculation of solvent entropies, either using total potential energy averages or van't Ho. plots, are carried out for the case of ion solvation in water. Although convergence problems will persist for large and complex simulation systems, good agreement with experiment is obtained here for relative and absolute ion hydration entropies. We also outline how solvent and solute entropic contributions are taken into account in empirical binding free energy calculations using the linear interaction energy method. In particular it is shown that empirical scaling of the nonpolar intermolecular ligand interaction energy effectively takes into account size dependent contributions to the binding free energy.

  • 26.
    Castell, Alina
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Fighting Tuberculosis –: Structural Studies of Three Mycobacterial Proteins2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis presents the cloning, purification, crystallization, and structural studies of two unknown proteins from Mycobacterium tuberculosis, and of an aminotransferase from Mycobacterium smegmatis. Structural knowledge of these proteins is of highest interest for structure-based drug design, which is one of the approaches that can be used in order to fight tuberculosis (TB).

    The structure of the conserved hypothetical protein Rv0216 was refined to a resolution of 1.9 Å. The structure exhibits a so-called double hotdog-fold, similar to known hydratases. However, only parts of the hydratase active site are conserved in Rv0216, and no function could be assigned to the protein. Several Rv0216-like protein sequences were found in a variety of actino- and proteobacteria, suggesting that these proteins form a new protein family. Furthermore, other hotdog-folded proteins in M. tuberculosis were identified, of which a few are likely to be hydratases or dehydratases involved in the fatty acid metabolism.

    The structure of Rv0130 exhibits a single hotdog-fold and contains a highly conserved R-hydratase motif. Rv0130 was shown to hydrate fatty acid coenzyme A derivatives with a length of six to eight carbons. The Rv0130 active site is situated in a long tunnel, formed by a kink in the central hotdog-helix, which indicate that it can utilize long fatty acid chains as well. A number of previously predicted hotdog-folded proteins also feature a similar tunnel.

    The structure of branched chain aminotransferase (BCAT) of M. smegmatis was determined in the apo-form and in complex with an aminooxy inhibitor. Mycobacterial BCAT is very similar to the human BCAT, apart for one important difference in the active site. Gly243 is a threonine in the human BCAT, a difference that offers specificity in inhibition and substrate recognition of these proteins. The aminooxy compound and MES were found to inhibit the mycobacterial BCAT activities. The aminooxy compound inhibits by blocking the substrate-pocket. A second inhibitor-binding site was identified through the binding of a MES molecule. Therefore, both the MES-binding site and the substrate-pocket of M. smegmatis BCAT are suggested to be potential sites for the development of new inhibitors against tuberculosis.

    List of papers
    1. Rv0216, a conserved hypothetical protein from Mycobacterium tuberculosis that is essential for bacterial
    Open this publication in new window or tab >>Rv0216, a conserved hypothetical protein from Mycobacterium tuberculosis that is essential for bacterial
    Show others...
    Manuscript (Other academic)
    Identifiers
    urn:nbn:se:uu:diva-97709 (URN)
    Available from: 2008-11-06 Created: 2008-11-06 Last updated: 2010-01-13Bibliographically approved
    2. Structure and function of Rv0130, a conserved hypothetical protein from Mycobacterium tuberculosis
    Open this publication in new window or tab >>Structure and function of Rv0130, a conserved hypothetical protein from Mycobacterium tuberculosis
    2006 (English)In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 15, no 10, p. 2300-2309Article in journal (Refereed) Published
    Abstract [en]

    A large fraction of the Mycobacterium tuberculosis genome codes for proteins of unknown function. We here report the structure of one of these proteins, Rv0130, solved to a resolution of 1.8 angstrom. The Rv0130 monomer features a single hotdog fold composed of a highly curved beta-sheet on top of a long and a short alpha-helix. Two monomers in turn pack to form a double-hotdog-folded homodimer, similar to a large group of enzymes that use thiol esters as substrates. Rv0130 was found to contain a highly conserved R-specific hydratase motif buried deeply between the two monomers. Our biochemical studies show that the protein is able to hydrate a short trans-2-enoyl-coenzyme A moiety with a k(cat) of 1.1 x 10(2) sec(-1). The importance of the side chains of D40 and H45 for hydratase activity is demonstrated by site-directed mutagenesis. In contrast to many hotdog-folded proteins, a proline residue distorts the central helix of Rv0130. This distortion allows the creation of a long, curved tunnel, similar to the substrate-binding channels of long-chain eukaryotic hydratase 2 enzymes.

    Keywords
    Rv0130, Mycobacterium tuberculosis, hydratase, hotdog fold, crystal structure
    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-94234 (URN)10.1110/ps.062309306 (DOI)000240851300008 ()16963641 (PubMedID)
    Available from: 2006-04-12 Created: 2006-04-12 Last updated: 2017-12-14Bibliographically approved
    3. Structural analysis of
    Open this publication in new window or tab >>Structural analysis of
    Manuscript (Other academic)
    Identifiers
    urn:nbn:se:uu:diva-97711 (URN)
    Available from: 2008-11-06 Created: 2008-11-06 Last updated: 2010-01-13Bibliographically approved
  • 27. Castell, Alina
    et al.
    Johansson, Patrik
    Unge, Torsten
    Jones, T Alwyn
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. Structural Biology Program.
    Bäckbro, Kristina
    Rv0216, a conserved hypothetical protein from Mycobacterium tuberculosis that2005In: Protein Sci, ISSN 0961-8368, Vol. 14, no 7, p. 1850-62Article in journal (Refereed)
  • 28. Chapman, M. S.
    et al.
    Liljas, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Structural Folds of Viral Proteins2003In: Advances in Protein Chemistry, ISSN 0065-3233, E-ISSN 1557-8941, Vol. 64, p. 125-196Article in journal (Other (popular science, discussion, etc.))
  • 29.
    Chaudhuri, BN
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, GJ
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Bjorkman, J
    Lehman-McKeeman, LD
    Oliver, JD
    Jones, TA
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    The structures of alpha(2u)-globulin and its complex with a hyaline droplet inducer1999In: ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, ISSN 0907-4449, Vol. 55, p. 753-762Article in journal (Refereed)
    Abstract [en]

    Alpha 2u-globulin (A2U) is the major urinary protein excreted by adult male rats. The structure of a monoclinic crystal form of A2U was reported in 1992 [Bocskei et al. (1992). Nature (London), 360, 186-188]. The structures of an orthorhombic crystal form of A2U at 2. 5 A resolution (refined to an R factor of 0.248; Rfree = 0.264) and of a complex between A2U and d-limonene 1,2-epoxide (DLO) at 2.9 A resolution (R factor = 0.248; Rfree = 0.260) are presented here. DLO is one of a diverse group of chemicals which cause a male rat-specific renal carcinogenesis called hyaline-droplet nephropathy. The rate-determining step in the development of this disorder is the binding of the toxin to A2U. Comparison of the cavities in A2U and in the corresponding mouse urinary protein (MUP) reveal that the former is tailor-made for small oval hydrophobic ligands such as DLO. The cavity in MUP is more shallow and elongated and cannot easily accommodate such ligands.

  • 30.
    Chaudhuri, BN
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, GJ
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Broutin-L'Hermite, I
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Bergfors, T
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Senn, H
    Le, Motte P
    Partouche, O
    Jones, TA
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Structures of cellular retinoic acid binding proteins I and II in complex with synthetic retinoids1999In: ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, ISSN 0907-4449, Vol. 55, p. 1850-1857Article in journal (Refereed)
    Abstract [en]

    Retinoids play important roles in diverse cellular processes including growth, cell differentiation and vision. Many natural and synthetic retinoids are used as drugs in dermatology and oncology. A large amount of data has been accumulated on the cellular activity of different synthetic retinoids. They are stabilized and transported inside the cell cytoplasm by binding and transport proteins, such as cellular retinol-binding proteins and cellular retinoic acid binding proteins (CRABPs). The structures of human CRABP II in complex with two different synthetic retinoids, Ro13-6307 and Ro12--7310 (at 2.1 and 2.0 A resolution, respectively) and of bovine CRABP I in complex with a retinobenzoic acid, Am80 (at 2.8 A resolution) are described. The binding affinities of human CRABP I and II for the retinoids studied here have been determined. All these compounds have comparable binding affinities (nanomolar range) for both CRABPs. Apart from the particular interactions of the carboxylate group of the retinoids with specific protein groups, each structure reveals characteristic interactions. Studying the atomic details of the interaction of retinoids with retinoid-binding proteins facilitates the understanding of the kinetics of retinoid trafficking inside the cytoplasm.

  • 31. Chen, Y W
    et al.
    Dodson, E J
    Kleywegt, G J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Does NMR mean 'Not for Molecular Replacement' ? Using NMR-based search models to solve protein crystal structures2000In: Structure, Vol. 8, p. R213-R220Article, review/survey (Other (popular scientific, debate etc.))
  • 32. Cohen, Serge X.
    et al.
    Morris, Richard J.
    Fernandez, Francisco J.
    Ben Jelloul, Marouane
    Kakaris, Mattheos
    Parthasarathy, Venkataraman
    Lamzin, Victor S
    Kleywegt, Gerard J.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Perrakis, Anastassis
    Towards complete validated models in the next generation of ARP/wARP2004In: Acta Crystallographica Section D: Biological Crystallography, ISSN 0907-4449, E-ISSN 1399-0047, Vol. 60, no Pt 12 Pt 1, p. 2222-9Article in journal (Refereed)
    Abstract [en]

    The design of a new versatile control system that will underlie future releases of the automated model-building package ARP/wARP is presented. A sophisticated expert system is under development that will transform ARP/wARP from a very useful model-building aid to a truly automated package capable of delivering complete, well refined and validated models comparable in quality to the result of intensive manual checking, rebuilding, hypothesis testing, refinement and validation cycles of an experienced crystallographer. In addition to the presentation of this control system, recent advances, ideas and future plans for improving the current model-building algorithms, especially for completing partially built models, are presented. Furthermore, a concept for integrating validation routines into the iterative model-building process is also presented.

  • 33. Coutard, B
    et al.
    Gorbalenya, A E
    Snijder, E J
    Leontovich, A M
    Poupon, A
    De Lamballerie, X
    Charrel, R
    Gould, E A
    Gunther, S
    Norder, H
    Klempa, B
    Bourhy, H
    Rohayem, J
    L'hermite, E
    Nordlund, P
    Stuart, D I
    Owens, R J
    Grimes, J M
    Tucker, P A
    Bolognesi, M
    Mattevi, A
    Coll, M
    Jones, T A
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, J
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Unge, T
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Hilgenfeld, R
    Bricogne, G
    Neyts, J
    La Colla, P
    Puerstinger, G
    Gonzalez, J P
    Leroy, E
    Cambillau, C
    Romette, J L
    Canard, B
    The VIZIER project: preparedness against pathogenic RNA viruses2008In: Antiviral Research, ISSN 0166-3542, E-ISSN 1872-9096, Vol. 78, no 1, p. 37-46Article in journal (Refereed)
    Abstract [en]

    Life-threatening RNA viruses emerge regularly, and often in an unpredictable manner. Yet, the very few drugs available against known RNA viruses have sometimes required decades of research for development. Can we generate preparedness for outbreaks of the, as yet, unknown viruses? The VIZIER (VIral enZymes InvolvEd in Replication) (http://www.vizier-europe.org/) project has been set-up to develop the scientific foundations for countering this challenge to society. VIZIER studies the most conserved viral enzymes (that of the replication machinery, or replicases) that constitute attractive targets for drug-design. The aim of VIZIER is to determine as many replicase crystal structures as possible from a carefully selected list of viruses in order to comprehensively cover the diversity of the RNA virus universe, and generate critical knowledge that could be efficiently utilized to jump-start research on any emerging RNA virus. VIZIER is a multidisciplinary project involving (i) bioinformatics to define functional domains, (ii) viral genomics to increase the number of characterized viral genomes and prepare defined targets, (iii) proteomics to express, purify, and characterize targets, (iv) structural biology to solve their crystal structures, and (v) pre-lead discovery to propose active scaffolds of antiviral molecules.

  • 34. Covarrubias, Adrian Suarez
    et al.
    Bergfors, Terese
    Jones, T Alwyn
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. Structural Biology Program.
    Högbom, Martin
    Structural mechanics of the pH-dependent activity of beta-carbonic anhydrase2006In: J Biol Chem, ISSN 0021-9258, Vol. 281, no 8, p. 4993-9Article in journal (Refereed)
  • 35.
    Covarrubias, Adrian Suarez
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Högbom, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Carroll, Paul
    Mannerstedt, Karin
    Oscarson, Stefan
    Parish, Tanya
    Jones, T. Alwyn
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Mowbray, Sherry
    Structural, biochemical and in vivo investigations of the threonine synthase from Mycobacterium tuberculosis2008In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 381, no 3, p. 622-633Article in journal (Refereed)
    Abstract [en]

    Threonine biosynthesis is a general feature of prokaryotes, eukaryotic microorganisms, and higher plants. Since mammals lack the appropriate synthetic machinery, instead obtaining the amino acid through their diet, the pathway is a potential focus for the development of novel antibiotics, antifungal agents, and herbicides. Threonine synthase (TS), a pyridoxal-5-phosphate-dependent enzyme, catalyzes the final step in the pathway, in which L-homoserine phosphate and water are converted into threonine and inorganic phosphate. In the present publication, we report structural and functional studies of Mycobacterium tuberculosis TS, the product of the rv1295 (thrC) gene. The structure gives new insights into the catalytic mechanism of TSs in general, specifically by suggesting the direct involvement of the phosphate moiety of the cofactor, rather than the inorganic phosphate product, in transferring a proton from C4' to C-gamma in the formation of the alpha beta-unsaturated aldimine. It further provides a basis for understanding why this enzyme has a higher pH optimum than has been reported elsewhere for TSs and gives rise to the prediction that the equivalent enzyme from Thermus thermophilus will exhibit similar behavior. A deletion of the relevant gene generated a strain of M. tuberculosis that requires threonine for growth, such auxotrophic strains are frequently attenuated in vivo, indicating that TS is a potential drug target in this organism.

  • 36. Davis, A M
    et al.
    Teague, S J
    Kleywegt, G J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Anwendung und Grenzen kristallographischer Daten im strukturbezogenen Liganden- und Wirkstoff-Design2003In: Angewandte Chemie, Vol. 115, p. 2822-2841Article, review/survey (Other (popular scientific, debate etc.))
  • 37. Davis, Andrew M.
    et al.
    St-Gallay, Stephen A.
    Kleywegt, Gerard J.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Limitations and lessons in the use of X-ray structural information in drug design2008In: Drug Discovery Today, ISSN 1359-6446, E-ISSN 1878-5832, Vol. 13, no 19-20, p. 831-841Article, review/survey (Refereed)
    Abstract [en]

    The use of X-ray crystal structure models continues to provide a strong stimulus to drug discovery, through the direct visualisation of ligand-receptor interactions. There is sometimes a limited appreciation of the uncertainties introduced during the process of deriving an atomic model from the experimentally observed electron density. Here, some of these uncertainties are highlighted with recent examples from the literature, together with snippets of advice for the medicinal chemist embarking on using X-ray crystal structure information in a drug discovery programme.

  • 38. Davis, Andrew M
    et al.
    Teague, Simon J
    Kleywegt, Gerard J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Application and limitations of X-ray crystallographic data in structure-based ligand and drug design.2003In: Angew Chem Int Ed Engl, ISSN 0570-0833, Vol. 42, no 24, p. 2718-36Article, review/survey (Other (popular scientific, debate etc.))
    Abstract [en]

    Structure-based design usually focuses upon the optimization of ligand affinity. However, successful drug design also requires the optimization of many other properties. The primary source of structural information for protein-ligand complexes is X-ray crystallography. The uncertainties introduced during the derivation of an atomic model from the experimentally observed electron density data are not always appreciated. Uncertainties in the atomic model can have significant consequences when this model is subsequently used as the basis of manual design, docking, scoring, and virtual screening efforts. Docking and scoring algorithms are currently imperfect. A good correlation between observed and calculated binding affinities is usually only observed only when very large ranges of affinity are considered. Errors in the correlation often exceed the range of affinities commonly encountered during lead optimization. Some structure-based design approaches now involve screening libraries by using technologies based on NMR spectroscopy and X-ray crystallography to discover small polar templates, which are used for further optimization. Such compounds are defined as leadlike and are also sought by more traditional high-throughput screening technologies. Structure-based design and HTS technologies show important complementarity and a degree of convergence.

  • 39.
    Dodson, E
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kleywegt, G J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Wilson, K
    Report of a workshop on the use of statistical validators in protein X-ray crystallography.1996In: Acta Crystallogr D Biol Crystallogr, ISSN 0907-4449, Vol. 52, no Pt 1, p. 228-34Article, review/survey (Other (popular scientific, debate etc.))
  • 40.
    Ekegren, Jenny K
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Unge, Torsten
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Safa, Mayada Zreik
    Wallberg, Hans
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Samuelsson, Bertil
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Hallberg, Anders
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    A new class of HIV-1 protease inhibitors containing a tertiary alcohol in the transition-state mimicking scaffold2005In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 48, no 25, p. 8098-8102Article in journal (Refereed)
    Abstract [en]

    Novel HIV-1 protease inhibitors encompassing a tertiary alcohol as part of the transition-state mimicking unit have been synthesized. Variation of the P1‘−P3‘ residues and alteration of the tertiary alcohol absolute stereochemistry afforded 10 inhibitors. High potencies for the compounds with (S)-configuration at the carbon carrying the tertiary hydroxyl group were achieved with Ki values down to 2.4 nM. X-ray crystallographic data for a representative compound in complex with HIV-1 protease are presented.

  • 41.
    Ericsson, Daniel
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Tunnels and Grooves: Structure-Function Studies in Two Disparate Enzymes2009Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis describes structural and binding studies in enzymes from two different  organisms: ribonucleotide reductase from Mycobacterium tuberculosis (RNR) and lipase A from Candida antarctica (CalA).

    RNR is viable as a target for new drugs against the causative agent of tuberculosis. The biologically active form of RNR is a heterotetramer with an α2β2 substructure. Here we show that an N-acetylated heptapeptide based on the C-terminal sequence of the smaller RNR subunit can disrupt the formation of the holoenzyme sufficiently to inhibit its function. An N-terminal truncation, an alanine scan and a novel statistical molecular design approach based on the heptapeptide Ac-Glu-Asp-Asp-Asp-Trp-Asp-Phe-OH were applied. A full-length acetylated heptapeptide was necessary for inhibition, and Trp5 and Phe7 were also essential. Exchanging the acetyl for the N-terminal Fmoc protective-group increased the binding potency ten-fold. Based on this, several truncated and N-protected peptides were evaluated in a competitive fluorescence polarization assay. The single-amino acid Fmoc-Trp inhibits the RNR holoenzyme formation with a dissociation constant of 12µM, making it an attractive candidate for further development of non-peptidic inhibitors

    Lipases are enzymes with major biotechnological applications. We report the x-ray structure of CalA, the first member of a novel family of lipases. The fold includes a well-defined lid as well as a classical α/β hydrolase domain. The structure is that of the closed/inactive state of the enzyme, but loop movements near Phe431 will provide virtually unlimited access to solvent for the alcohol moiety of an ester substrate. The structure thus provides a basis for understanding the enzyme's preference for acyl moieties with long, straight tails, and for its highly promiscuous acceptance of widely different alcohol and amine moieties. An unconventional oxyanion hole is observed in the present structure, although the situation may change during interfacial activation.

    List of papers
    1. Design, synthesis and evaluation of peptide inhibitors of Mycobacterium tuberculosis ribonucleotide reductase
    Open this publication in new window or tab >>Design, synthesis and evaluation of peptide inhibitors of Mycobacterium tuberculosis ribonucleotide reductase
    Show others...
    2007 (English)In: Journal of Peptide Science, ISSN 1075-2617, E-ISSN 1099-1387, Vol. 13, no 12, p. 822-832Article in journal (Refereed) Published
    Abstract [en]

    Mycobacterium tuberculosis ribonucleotide reductase (RNR) is a potential target for new antitubercular drugs. Herein we describe the synthesis and evaluation of peptide inhibitors of RNR derived from the C-terminus of the small subunit of M. tuberculosis RNR. An N-terminal truncation, an alanine scan and a novel statistical molecular design (SMD) approach based on the heptapeptide Ac-Glu-Asp-Asp-Asp-Trp-Asp-Phe-OH were applied in this study. The alanine scan showed that TrP5 and Phe7 were important for inhibitory potency. A quantitative structure relationship (QSAR) model was developed based on the synthesized peptides which showed that a negative charge in positions 2, 3, and 6 is beneficial for inhibitory potency. Finally, in position 5 the model coefficients indicate that there is room for a larger side chain., as compared to Trp5.

    Keywords
    mycobacterium tuberculosis, ribonucleotide reductase, peptide inhibitors, alanine scan, statistical molecular design, structure activity relationships, FHDoE
    National Category
    Pharmaceutical Sciences
    Identifiers
    urn:nbn:se:uu:diva-14261 (URN)10.1002/psc.906 (DOI)000252000600007 ()17918768 (PubMedID)
    Available from: 2008-05-29 Created: 2008-05-29 Last updated: 2018-01-12Bibliographically approved
    2. Identification of small peptides mimicking the R2 C-terminus of Mycobacterium tuberculosis ribonucleotide reductase
    Open this publication in new window or tab >>Identification of small peptides mimicking the R2 C-terminus of Mycobacterium tuberculosis ribonucleotide reductase
    Show others...
    2010 (English)In: Journal of Peptide Science, ISSN 1075-2617, E-ISSN 1099-1387, Vol. 16, no 3, p. 159-164Article in journal (Refereed) Published
    Abstract [en]

    Ribonucleotide reductase (RNR) is a viable target for new drugs against the causative agent of tuberculosis, Mycobacterium tuberculosis. Previous work has shown that an N-acetylated heptapeptide based on the C-terminal sequence of the smaller RNR subunit can disrupt the formation of the holoenzyme sufficiently to inhibit its function. Here the synthesis and binding affinity, evaluated by competitive fluorescence polarization, of several truncated and N-protected peptides are described. The protected single-amino acid Fmoc-Trp shows binding affinity comparable to the N-acetylated heptapeptide, making it an attractive candidate for further development of non-peptidic RNR inhibitors.

    Keywords
    Fluorescence polarization, Mycobacterium tuberculosis, Peptide inhibitors, Ribonucleotide reductase
    National Category
    Pharmaceutical Sciences
    Identifiers
    urn:nbn:se:uu:diva-112344 (URN)10.1002/psc.1214 (DOI)000275448300007 ()20127854 (PubMedID)
    Available from: 2010-01-26 Created: 2010-01-13 Last updated: 2018-01-12Bibliographically approved
    3. X-ray structure of Candida antarctica lipase A shows a novel lid structure and a likely mode of interfacial activation
    Open this publication in new window or tab >>X-ray structure of Candida antarctica lipase A shows a novel lid structure and a likely mode of interfacial activation
    Show others...
    2008 (English)In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 376, no 1, p. 109-119Article in journal (Refereed) Published
    Abstract [en]

    In nature, lipases (EC 3.1.1.3) catalyze the hydrolysis of triglycerides to form glycerol and fatty acids. Under the appropriate conditions, the reaction is reversible, and so biotechnological applications commonly make use of their capacity for esterification as well as for hydrolysis of a wide variety of compounds. In the present paper, we report the X-ray structure of lipase A from Candida antarctica, solved by single isomorphous replacement with anomalous scattering, and refined to 2.2-A resolution. The structure is the first from a novel family of lipases. Contrary to previous predictions, the fold includes a well-defined lid as well as a classic alpha/beta hydrolase domain. The catalytic triad is identified as Ser184, Asp334 and His366, which follow the sequential order considered to be characteristic of lipases; the serine lies within a typical nucleophilic elbow. Computer docking studies, as well as comparisons to related structures, place the carboxylate group of a fatty acid product near the serine nucleophile, with the long lipid tail closely following the path through the lid that is marked by a fortuitously bound molecule of polyethylene glycol. For an ester substrate to bind in an equivalent fashion, loop movements near Phe431 will be required, suggesting the primary focus of the conformational changes required for interfacial activation. Such movements will provide virtually unlimited access to solvent for the alcohol moiety of an ester substrate. The structure thus provides a basis for understanding the enzyme's preference for acyl moieties with long, straight tails, and for its highly promiscuous acceptance of widely different alcohol and amine moieties. An unconventional oxyanion hole is observed in the present structure, although the situation may change during interfacial activation.

    Keywords
    lipase, interfacial activation, hydrolase, X-ray structure, substrate specificity
    National Category
    Biochemistry and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-14221 (URN)10.1016/j.jmb.2007.10.079 (DOI)000253181500011 ()18155238 (PubMedID)
    Available from: 2008-01-29 Created: 2008-01-29 Last updated: 2017-12-11Bibliographically approved
  • 42.
    Ericsson, Daniel J.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Kasrayan, Alex
    Johansson, Patrik
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Bergfors, Terese
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Sandström, Anders G.
    Bäckvall, Jan-Erling
    Mowbray, Sherry L.
    Department of Molecular Biology, Swedish University of Agricultural Sciences, Biomedical Center, Uppsala, Sweden.
    X-ray structure of Candida antarctica lipase A shows a novel lid structure and a likely mode of interfacial activation2008In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 376, no 1, p. 109-119Article in journal (Refereed)
    Abstract [en]

    In nature, lipases (EC 3.1.1.3) catalyze the hydrolysis of triglycerides to form glycerol and fatty acids. Under the appropriate conditions, the reaction is reversible, and so biotechnological applications commonly make use of their capacity for esterification as well as for hydrolysis of a wide variety of compounds. In the present paper, we report the X-ray structure of lipase A from Candida antarctica, solved by single isomorphous replacement with anomalous scattering, and refined to 2.2-A resolution. The structure is the first from a novel family of lipases. Contrary to previous predictions, the fold includes a well-defined lid as well as a classic alpha/beta hydrolase domain. The catalytic triad is identified as Ser184, Asp334 and His366, which follow the sequential order considered to be characteristic of lipases; the serine lies within a typical nucleophilic elbow. Computer docking studies, as well as comparisons to related structures, place the carboxylate group of a fatty acid product near the serine nucleophile, with the long lipid tail closely following the path through the lid that is marked by a fortuitously bound molecule of polyethylene glycol. For an ester substrate to bind in an equivalent fashion, loop movements near Phe431 will be required, suggesting the primary focus of the conformational changes required for interfacial activation. Such movements will provide virtually unlimited access to solvent for the alcohol moiety of an ester substrate. The structure thus provides a basis for understanding the enzyme's preference for acyl moieties with long, straight tails, and for its highly promiscuous acceptance of widely different alcohol and amine moieties. An unconventional oxyanion hole is observed in the present structure, although the situation may change during interfacial activation.

  • 43.
    Ericsson, Daniel J.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Nurbo, Johanna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Muthas, Daniel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Hertzberg, Kalle
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Lindeberg, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Karlén, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Unge, Torsten
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Identification of small peptides mimicking the R2 C-terminus of Mycobacterium tuberculosis ribonucleotide reductase2010In: Journal of Peptide Science, ISSN 1075-2617, E-ISSN 1099-1387, Vol. 16, no 3, p. 159-164Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductase (RNR) is a viable target for new drugs against the causative agent of tuberculosis, Mycobacterium tuberculosis. Previous work has shown that an N-acetylated heptapeptide based on the C-terminal sequence of the smaller RNR subunit can disrupt the formation of the holoenzyme sufficiently to inhibit its function. Here the synthesis and binding affinity, evaluated by competitive fluorescence polarization, of several truncated and N-protected peptides are described. The protected single-amino acid Fmoc-Trp shows binding affinity comparable to the N-acetylated heptapeptide, making it an attractive candidate for further development of non-peptidic RNR inhibitors.

  • 44.
    Ersmark, Karolina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Nervall, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Gutiérrez-de-Terán, Hugo
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Hamelink, Elizabeth
    Janka, Linda K.
    Clemente, Jose C.
    Dunn, Ben M.
    Gogoll, Adolf
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Samuelsson, Bertil
    Åqvist, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Hallberg, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry I. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Macrocyclic inhibitors of the malarial aspartic proteases plasmepsin I, II, and IV2006In: Biorganic & Medicinal Chemistry, no 14, p. 2197-2208Article in journal (Refereed)
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    Ersmark, Karolina
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry. Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. ORGFARM.
    Nervall, Martin
    Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Hamelink, Elizabeth
    Janka, Linda K
    Clemente, Jose C
    Dunn, Ben M
    Blackman, Michael J
    Samuelsson, Bertil
    Aqvist, Johan
    Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Hallberg, Anders
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry. Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology. ORGFARM.
    Synthesis of malarial plasmepsin inhibitors and prediction of binding modes by molecular dynamics simulations.2005In: J Med Chem, ISSN 0022-2623, Vol. 48, no 19, p. 6090-6106Article in journal (Refereed)
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    Feierberg, I.
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Åqvist, J
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Molecular Biology.
    Correction: The Catalytic Power of Ketosteroid Isomerase Investigated by Computer Simulation2003In: Biochemistry, Vol. 42, p. 2258-Article in journal (Refereed)