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Andersson, Eva
Publications (9 of 9) Show all publications
Karlsson, E., Andersson, E., Dogan, J., Gianni, S., Jemth, P. & Camilloni, C. (2019). A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins. Journal of Biological Chemistry, 294(4), 1230-1239
Open this publication in new window or tab >>A structurally heterogeneous transition state underlies coupled binding and folding of disordered proteins
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2019 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 294, no 4, p. 1230-1239Article in journal (Refereed) Published
Abstract [en]

Many intrinsically disordered proteins (IDPs) attain a well-defined structure in a coupled folding and binding reaction with another protein. Such reactions may involve early to late formation of different native structural regions along the reaction pathway. To obtain insights into the transition state for a coupled binding and folding reaction, we performed restrained molecular dynamics simulations using previously determined experimental binding phi(b) values of the interaction between two IDP domains: the activation domain from the p160 transcriptional co-activator for thyroid hormone and retinoid receptors (ACTR) and the nuclear co-activator binding domain (NCBD) of CREB-binding protein, each forming three well-defined alpha-helices upon binding. These simulations revealed that both proteins are largely disordered in the transition state for complex formation, except for two helices, one from each domain, that display a native-like structure. The overall transition state structure was extended and largely dynamic with many weakly populated contacts. To test the transition state model, we combined site-directed mutagenesis with kinetic experiments, yielding results consistent with overall diffuse interactions and formation of native intramolecular interactions in the third NCBD helix during the binding reaction. Our findings support the view that the transition state and, by inference, any encounter complex in coupled binding and folding reactions are structurally heterogeneous and largely independent of specific interactions. Furthermore, experimental phi(b) values and Bronsted plots suggested that the transition state is globally robust with respect to most mutations but can display more native-like features for some highly destabilizing mutations, possibly because of Hammond behavior or ground-state effects.

Keywords
intrinsically disordered protein, pre-steady-state kinetics, protein folding, protein-protein interaction, molecular dynamics
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-377701 (URN)10.1074/jbc.RA118.005854 (DOI)000457879500014 ()30514761 (PubMedID)
Funder
Swedish Research Council, 2016-04965
Available from: 2019-02-25 Created: 2019-02-25 Last updated: 2019-02-25Bibliographically approved
Karlsson, E., Andersson, E., Jones, N. C., Hoffmann, S. V., Jemth, P. & Kjaergaard, M. (2019). Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism. Biophysical Journal, 117(4), 729-742
Open this publication in new window or tab >>Coupled Binding and Helix Formation Monitored by Synchrotron-Radiation Circular Dichroism
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2019 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 117, no 4, p. 729-742Article in journal (Refereed) Published
Abstract [en]

Intrinsically disordered proteins organize interaction networks in the cell in many regulation and signaling processes. These proteins often gain structure upon binding to their target proteins in multistep reactions involving the formation of both secondary and tertiary structure. To understand the interactions of disordered proteins, we need to understand the mechanisms of these coupled folding and binding reactions. We studied helix formation in the binding of the molten globule-like nuclear coactivator binding domain and the disordered interaction domain from activator of thyroid hormone and retinoid receptors. We demonstrate that helix formation in a rapid binding reaction can be followed by stopped-flow synchrotron-radiation circular dichroism (CD) spectroscopy and describe the design of such a beamline. Fluorescence-monitored binding experiments of activator of thyroid hormone and retinoid receptors and nuclear coactivator binding domain display several kinetic phases, including one concentration-independent phase, which is consistent with an intermediate stabilized at high ionic strength. Time-resolved CD experiments show that almost all helicity is formed upon initial association of the proteins or separated from the encounter complex by only a small energy barrier. Through simulation of mechanistic models, we show that the intermediate observed at high ionic strength likely involves a structural rearrangement with minor overall changes in helicity. Our experiments provide a benchmark for simulations of coupled binding reactions and demonstrate the feasibility of using synchrotron-radiation CD for mechanistic studies of protein-protein interactions.

Place, publisher, year, edition, pages
CELL PRESS, 2019
National Category
Biophysics
Identifiers
urn:nbn:se:uu:diva-393728 (URN)10.1016/j.bpj.2019.07.014 (DOI)000482097100011 ()31378314 (PubMedID)
Funder
Swedish Research Council, 2016-04965
Available from: 2019-09-27 Created: 2019-09-27 Last updated: 2019-09-27Bibliographically approved
Nyqvist, I., Andersson, E. & Dogan, J. (2019). Role of Conformational Entropy in Molecular Recognition by TAZ1 of CBP. Journal of Physical Chemistry B, 123(13), 2882-2888
Open this publication in new window or tab >>Role of Conformational Entropy in Molecular Recognition by TAZ1 of CBP
2019 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 123, no 13, p. 2882-2888Article in journal (Refereed) Published
Abstract [en]

The globular transcriptional adapter zinc binding 1 (TAZ1) domain of CREB binding protein participates in protein-protein interactions that are involved in transcriptional regulation. TAZ1 binds numerous targets, of which many are intrinsically disordered proteins that undergo a disorder-to-order transition to various degrees. One such target is the disordered transactivation domain of transcription factor RelA (TAD-RelA), which with its interaction with TAZ1 is involved in transcriptional regulation of genes in NF-kappa B signaling. We have here performed nuclear magnetic resonance backbone and side-chain relaxation studies to investigate the influence of RelA-TA2 (residues 425-490 in TAD-RelA) binding on the subnanosecond internal motions of TAZ1. We find a considerable dynamic response on both the backbone and side-chain levels, which corresponds to a conformational entropy change that contributes significantly to the binding energetics. We further show that the microscopic origins of the dynamic response of TAZ1 vary depending on the target. This study demonstrates that folded protein domains that are able to interact with various targets are not dynamically passive but can have a significant role in the motional response upon target association.

Place, publisher, year, edition, pages
AMER CHEMICAL SOC, 2019
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-382565 (URN)10.1021/acs.jpcb.9b01343 (DOI)000463844200015 ()30860376 (PubMedID)
Funder
Swedish Research CouncilCarl Tryggers foundation Åke Wiberg FoundationLars Hierta Memorial FoundationMagnus Bergvall Foundation
Available from: 2019-05-03 Created: 2019-05-03 Last updated: 2019-05-03Bibliographically approved
Åberg, E., Karlsson, O. A., Andersson, E. & Jemth, P. (2018). Binding Kinetics of the Intrinsically Disordered p53 Family Transactivation Domains and MDM2. Journal of Physical Chemistry B, 122(27), 6899-6905
Open this publication in new window or tab >>Binding Kinetics of the Intrinsically Disordered p53 Family Transactivation Domains and MDM2
2018 (English)In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 122, no 27, p. 6899-6905Article in journal (Refereed) Published
Abstract [en]

Because of their prominent roles in cell-cycle regulation and cancer, the interaction between MDM2 and the intrinsically disordered transactivation domain (TAD) of p53 is exceptionally well-studied. However, although there are numerous computational studies on the interaction mechanism, there is a paucity of experimental data regarding the kinetics and mechanism. We have used stopped flow fluorescence to investigate the binding reaction between MDM2 and TAD from p53 as well as from its paralogs p63 and p73, and in particular, focused on the salt dependence of the interaction. The observed kinetics are consistent with a two-state mechanism within the time frame of the stopped flow methodology; thus, any conformational changes including the previously identified MDM2 lid dynamics must occur on a time scale <5 ms at 10 °C. The association rate constants are similar for the three TADs, and differences in the dissociation rate constants determine the various affinities with MDM2. In contrast to previous studies, we found a relatively small ionic-strength dependence for all three interactions, highlighting the large variation in the role of electrostatics among binding reactions of intrinsically disordered proteins (IDPs). The basal association rate constants in the absence of electrostatic interactions were relatively high (≥2 × 106 M–1 s–1 at 10 °C), suggesting that a large number of initial contacts may lead to a productive complex. Our findings support an emerging picture of “conformational funneling” occurring in the initial stages of interactions involving IDPs and that these early binding events can rely on hydrophobic as well as charge–charge interactions.

National Category
Biochemistry and Molecular Biology Physical Chemistry
Identifiers
urn:nbn:se:uu:diva-361528 (URN)10.1021/acs.jpcb.8b03876 (DOI)000439002900006 ()29878773 (PubMedID)
Funder
Swedish Research Council, 2016-04134
Available from: 2018-09-25 Created: 2018-09-25 Last updated: 2018-09-25Bibliographically approved
Jemth, P., Karlsson, E., Vogeli, B., Guzovsky, B., Andersson, E., Hultqvist, G., . . . Chi, C. N. (2018). Structure and dynamics conspire in the evolution of affinity between intrinsically disordered proteins. Science Advances, 4(10), Article ID eaau4130.
Open this publication in new window or tab >>Structure and dynamics conspire in the evolution of affinity between intrinsically disordered proteins
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2018 (English)In: Science Advances, E-ISSN 2375-2548, Vol. 4, no 10, article id eaau4130Article in journal (Refereed) Published
Abstract [en]

In every established species, protein-protein interactions have evolved such that they are fit for purpose. However, the molecular details of the evolution of new protein-protein interactions are poorly understood. We have used nuclear magnetic resonance spectroscopy to investigate the changes in structure and dynamics during the evolution of a protein-protein interaction involving the intrinsically disordered CREBBP (CREB-binding protein) interaction domain (CID) and nuclear coactivator binding domain (NCBD) from the transcriptional coregulators NCOA (nuclear receptor coactivator) and CREBBP/p300, respectively. The most ancient low-affinity "Cambrian-like" [540 to 600 million years (Ma) ago] CID/NCBD complex contained less secondary structure and was more dynamic than the complexes from an evolutionarily younger "Ordovician-Silurian" fish ancestor (ca. 440 Ma ago) and extant human. The most ancient Cambrian-like CID/NCBD complex lacked one helix and several interdomain interactions, resulting in a larger solvent-accessible surface area. Furthermore, the most ancient complex had a high degree of millisecond-to-microsecond dynamics distributed along the entire sequences of both CID and NCBD. These motions were reduced in the Ordovician-Silurian CID/NCBD complex and further redistributed in the extant human CID/NCBD complex. Isothermal calorimetry experiments show that complex formation is enthalpically favorable and that affinity is modulated by a largely unfavorable entropic contribution to binding. Our data demonstrate how changes in structure and motion conspire to shape affinity during the evolution of a protein-protein complex and provide direct evidence for the role of structural, dynamic, and frustrational plasticity in the evolution of interactions between intrinsically disordered proteins.

Place, publisher, year, edition, pages
AMER ASSOC ADVANCEMENT SCIENCE, 2018
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-369756 (URN)10.1126/sciadv.aau4130 (DOI)000449221200069 ()30397651 (PubMedID)
Funder
Swedish Research Council
Available from: 2018-12-17 Created: 2018-12-17 Last updated: 2018-12-17Bibliographically approved
Lindstrom, I., Andersson, E. & Dogan, J. (2018). The transition state structure for binding between TAZ1 of CBP and the disordered Hif-1 alpha CAD. Scientific Reports, 8, Article ID 7872.
Open this publication in new window or tab >>The transition state structure for binding between TAZ1 of CBP and the disordered Hif-1 alpha CAD
2018 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, article id 7872Article in journal (Refereed) Published
Abstract [en]

Intrinsically disordered proteins (IDPs) are common in eukaryotes. However, relatively few experimental studies have addressed the nature of the rate-limiting transition state for the coupled binding and folding reactions involving IDPs. By using site-directed mutagenesis in combination with kinetics measurements we have here characterized the transition state for binding between the globular TAZ1 domain of CREB binding protein and the intrinsically disordered C-terminal activation domain of Hif-1 alpha (Hif-1 alpha CAD). A total of 17 Hif-1 alpha CAD point-mutations were generated and a F-value binding analysis was carried out. We found that native hydrophobic binding interactions are not formed at the transition state. We also investigated the effect the biologically important Hif-1 alpha CAD Asn-803 hydroxylation has on the binding kinetics, and found that the whole destabilization effect due the hydroxylation is within the dissociation rate constant. Thus, the rate-limiting transition state is "disordered-like", with native hydrophobic binding contacts being formed cooperatively after the rate-limiting barrier, which is clearly shown by linear free energy relationships. The same behavior was observed in a previously characterized TAZ1/IDP interaction, which may suggest common features for the rate-limiting transition state for TAZ1/IDP interactions.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2018
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-357283 (URN)10.1038/s41598-018-26213-x (DOI)000432441400060 ()29777197 (PubMedID)
Funder
Swedish Research CouncilCarl Tryggers foundation Magnus Bergvall FoundationÅke Wiberg Foundation
Available from: 2018-08-14 Created: 2018-08-14 Last updated: 2018-08-14Bibliographically approved
Hultqvist, G., Åberg, E., Camilloni, C., Sundell, G., Andersson, E., Dogan, J., . . . Jemth, P. (2017). Emergence and evolution of an interaction between intrinsically disordered proteins. eLIFE, 6, Article ID e16059.
Open this publication in new window or tab >>Emergence and evolution of an interaction between intrinsically disordered proteins
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2017 (English)In: eLIFE, E-ISSN 2050-084X, Vol. 6, article id e16059Article in journal (Refereed) Published
Abstract [en]

Protein-protein interactions involving intrinsically disordered proteins are important for cellular function and common in all organisms. However, it is not clear how such interactions emerge and evolve on a molecular level. We performed phylogenetic reconstruction, resurrection and biophysical characterization of two interacting disordered protein domains, CID and NCBD. CID appeared after the divergence of protostomes and deuterostomes 450-600 million years ago, while NCBD was present in the protostome/deuterostome ancestor. The most ancient CID/NCBD formed a relatively weak complex (K(d similar to)5 mu M). At the time of the first vertebrate-specific whole genome duplication, the affinity had increased (K-d\similar to 200 nM) and was maintained in further speciation. Experiments together with molecular modeling using NMR chemical shifts suggest that new interactions involving intrinsically disordered proteins may evolve via a low-affinity complex which is optimized by modulating direct interactions as well as dynamics, while tolerating several potentially disruptive mutations.

National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-322818 (URN)10.7554/eLife.16059 (DOI)000400663100001 ()
Available from: 2017-09-12 Created: 2017-09-12 Last updated: 2018-01-28Bibliographically approved
Toto, A., Pedersen, S. W., Karlsson, O. A., Moran, G. E., Andersson, E., Chi, C. N., . . . Jemth, P. (2016). Ligand binding to the PDZ domains of postsynaptic density protein 95. Protein Engineering Design & Selection, 29(5), 169-175
Open this publication in new window or tab >>Ligand binding to the PDZ domains of postsynaptic density protein 95
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2016 (English)In: Protein Engineering Design & Selection, ISSN 1741-0126, E-ISSN 1741-0134, Vol. 29, no 5, p. 169-175Article in journal (Refereed) Published
Abstract [en]

Cellular scaffolding and signalling is generally governed by multidomain proteins, where each domain has a particular function. Postsynaptic density protein 95 (PSD-95) is involved in synapse formation and is a typical example of such a multidomain protein. Protein-protein interactions of PSD-95 are well studied and include the following three protein ligands: (i) N-methyl-d-aspartate-type ionotropic glutamate receptor subunit GluN2B, (ii) neuronal nitric oxide synthase and (iii) cysteine-rich protein (CRIPT), all of which bind to one or more of the three PDZ domains in PSD-95. While interactions for individual PDZ domains of PSD-95 have been well studied, less is known about the influence of neighbouring domains on the function of the respective individual domain. We therefore performed a systematic study on the ligand-binding kinetics of PSD-95 using constructs of different size for PSD-95 and its ligands. Regarding the canonical peptide-binding pocket and relatively short peptides (up to 15-mer), the PDZ domains in PSD-95 by and large work as individual binding modules. However, in agreement with previous studies, residues outside of the canonical binding pocket modulate the affinity of the ligands. In particular, the dissociation of the 101 amino acid CRIPT from PSD-95 is slowed down at least 10-fold for full-length PSD-95 when compared with the individual PDZ3 domain.

Keywords
CRIPT, GluN2B, Kinetics, PDZ domain, PSD-95
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-298245 (URN)10.1093/protein/gzw004 (DOI)000376351600002 ()26941280 (PubMedID)
Funder
Swedish Research Council, 2012-5096
Available from: 2016-07-01 Created: 2016-07-01 Last updated: 2017-11-28Bibliographically approved
Dogan, J., Jonasson, J., Andersson, E. & Jemth, P. (2015). Binding Rate Constants Reveal Distinct Features of Disordered Protein Domains. Biochemistry, 54(30), 4741-4750
Open this publication in new window or tab >>Binding Rate Constants Reveal Distinct Features of Disordered Protein Domains
2015 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 54, no 30, p. 4741-4750Article in journal (Refereed) Published
Abstract [en]

Intrinsically disordered proteins (IDPs) are abundant in the proteome and involved in key cellular functions. However, experimental data about the binding kinetics of IDPs as a function of different environmental conditions are scarce. We have performed an extensive characterization of the ionic strength dependence of the interaction between the molten globular nuclear co-activator binding domain (NCBD) of CREB binding protein and five different protein ligands, including the intrinsically disordered activation domain of p160 transcriptional co-activators (SRC1, TIF2, ACTR), the p53 transactivation domain, and the folded pointed domain (PNT) of transcription factor ETS-2. Direct comparisons of the binding rate constants under identical conditions show that the association rate constant, k(on), for interactions between NCBD and disordered protein domains is high at low salt concentrations (90-350 x 10(6) M-1 s(-1) at 4 degrees C) but is reduced significantly (10-30-fold) with an increasing ionic strength and reaches a plateau around physiological ionic strength. In contrast, the k(on) for the interaction between NCBD and the folded PNT domain is only 7 x 10(6) M-1 s(-1) (4 degrees C and low salt) and displays weak ionic strength dependence, which could reflect a distinctly different association that relies less on electrostatic interactions. Furthermore, the basal rate constant (in the absence of electrostatic interactions) is high for the NCBD interactions, exceeding those typically observed for folded proteins. One likely interpretation is that disordered proteins have a large number of possible collisions leading to a productive on-pathway encounter complex, while folded proteins are more restricted in terms of orientation. Our results highlight the importance of electrostatic interactions in binding involving IDPs and emphasize the significance of including ionic strength as a factor in studies that compare the binding properties of IDPs to those of ordered proteins.

National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-261964 (URN)10.1021/acs.biochem.5b00520 (DOI)000359277800016 ()26153298 (PubMedID)
Funder
Swedish Research Council
Available from: 2015-09-09 Created: 2015-09-07 Last updated: 2017-12-04Bibliographically approved
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