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Jemth, Per
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Publications (10 of 81) Show all publications
Gautier, C., Visconti, L., Jemth, P. & Gianni, S. (2017). Addressing the role of the alpha-helical extension in the folding of the third PDZ domain from PSD-95. Scientific Reports, 7, Article ID 12593.
Open this publication in new window or tab >>Addressing the role of the alpha-helical extension in the folding of the third PDZ domain from PSD-95
2017 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 7, article id 12593Article in journal (Refereed) Published
Abstract [en]

PDZ domains are one of the most important protein-protein interaction domains in human. While presenting a conserved three dimensional structure, a substantial number of PDZ domains display structural extensions suggested to be involved in their folding and binding mechanisms. The C-terminal a-helix extension (alpha 3) of the third PDZ domain from PSD-95 (PDZ3) has been reported to have a role in function of the domain as well as in the stabilization of the native fold. Here we report an evaluation of the effect of the truncation of this additional helix on the folding and unfolding kinetics of PDZ3. Fluorescent variants of full length and truncated PDZ3 were produced and stopped-flow fluorescence measurements were made under different experimental conditions (pH, ionic strength and temperature) to investigate the folding kinetics of the respective variant. The results show that folding of PDZ3 is robust and that the mechanism is only marginally affected by the truncation, which contributes to a destabilization of the native state, but otherwise do not change the overall observed kinetics. Furthermore, the increase in the unfolding rate constants, but not the folding rate constant upon deletion of alpha 3 suggests that the a-helical extension is largely unstructured in the folding transition state.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2017
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-337114 (URN)10.1038/s41598-017-12827-0 (DOI)000412138800029 ()28974728 (PubMedID)
Funder
EU, Horizon 2020, 675341
Available from: 2017-12-21 Created: 2017-12-21 Last updated: 2017-12-21Bibliographically 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
Åberg, E., Saccoccia, F., Grabherr, M., Ore, W. Y., Jemth, P. & Hultqvist, G. (2017). Evolution of the p53-MDM2 pathway. BMC Evolutionary Biology, 17, Article ID 177.
Open this publication in new window or tab >>Evolution of the p53-MDM2 pathway
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2017 (English)In: BMC Evolutionary Biology, ISSN 1471-2148, E-ISSN 1471-2148, Vol. 17, article id 177Article in journal (Refereed) Published
Abstract [en]

Background: The p53 signalling pathway, which controls cell fate, has been extensively studied due to its prominent role in tumor development. The pathway includes the tumor supressor protein p53, its vertebrate paralogs p63 and p73, and their negative regulators MDM2 and MDM4. The p53/p63/p73-MDM system is ancient and can be traced in all extant animal phyla. Despite this, correct phylogenetic trees including both vertebrate and invertebrate species of the p53/p63/p73 and MDM families have not been published. Results: Here, we have examined the evolution of the p53/p63/p73 protein family with particular focus on the p53/ p63/p73 transactivation domain (TAD) and its co-evolution with the p53/p63/p73- binding domain (p53/p63/p73BD) of MDM2. We found that the TAD and p53/p63/p73BD share a strong evolutionary connection. If one of the domains of the protein is lost in a phylum, then it seems very likely to be followed by loss of function by the other domain as well, and due to the loss of function it is likely to eventually disappear. By focusing our phylogenetic analysis to p53/p63/ p73 and MDM proteins from phyla that retain the interaction domains TAD and p53/p63/p73BD, we built phylogenetic trees of p53/p63/p73 and MDM based on both vertebrate and invertebrate species. The trees follow species evolution and contain a total number of 183 and 98 species for p53/p63/p73 and MDM, respectively. We also demonstrate that the p53/p63/p73 and MDM families result from whole genome duplications. Conclusions: The signaling pathway of the TAD and p53/p63/p73BD in p53/p63/p73 and MDM, respectively, dates back to early metazoan time and has since then tightly co-evolved, or disappeared in distinct lineages.

Place, publisher, year, edition, pages
BioMed Central, 2017
Keywords
p53, MDM, Co-evolution, Phylogeny
National Category
Evolutionary Biology
Identifiers
urn:nbn:se:uu:diva-334047 (URN)10.1186/s12862-017-1023-y (DOI)000407013500001 ()28774266 (PubMedID)
Available from: 2017-11-21 Created: 2017-11-21 Last updated: 2018-01-28Bibliographically approved
Gianni, S. & Jemth, P. (2017). How Fast Is Protein-Ligand Association?. TIBS -Trends in Biochemical Sciences. Regular ed., 42(11), 847-849
Open this publication in new window or tab >>How Fast Is Protein-Ligand Association?
2017 (English)In: TIBS -Trends in Biochemical Sciences. Regular ed., ISSN 0968-0004, E-ISSN 1362-4326, Vol. 42, no 11, p. 847-849Article in journal, Editorial material (Other academic) Published
Abstract [en]

There is increasing interest in studying protein interactions and their role in cell biology using kinetics. However, there is confusion about the proper terminology in terms of the distinction between rates and rate constants. We recommend a more stringent use of the words speed, fast, slow, rate, and rate constant.

National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-346754 (URN)10.1016/j.tibs.2017.08.007 (DOI)000413441400002 ()
Available from: 2018-03-27 Created: 2018-03-27 Last updated: 2018-03-27Bibliographically approved
Thalmann, D. S., Ring, H., Sundström, E., Cao, X., Larsson, M., Kerje, S., . . . Andersson, L. (2017). The evolution of Sex-linked barring alleles in chickens involves both regulatory and coding changes in CDKN2A. PLoS Genetics, 13(4), Article ID e1006665.
Open this publication in new window or tab >>The evolution of Sex-linked barring alleles in chickens involves both regulatory and coding changes in CDKN2A
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2017 (English)In: PLoS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 13, no 4, article id e1006665Article in journal (Refereed) Published
Abstract [en]

Sex-linked barring is a fascinating plumage pattern in chickens recently shown to be associated with two non-coding and two missense mutations affecting the ARF transcript at the CDKN2A tumor suppressor locus. It however remained a mystery whether all four mutations are indeed causative and how they contribute to the barring phenotype. Here, we show that Sex-linked barring is genetically heterogeneous, and that the mutations form three functionally different variant alleles. The B0 allele carries only the two non-coding changes and is associated with the most dilute barring pattern, whereas the B1 and B2 alleles carry both the two non-coding changes and one each of the two missense mutations causing the Sex-linked barring and Sex-linked dilution phenotypes, respectively. The data are consistent with evolution of alleles where the non-coding changes occurred first followed by the two missense mutations that resulted in a phenotype more appealing to humans. We show that one or both of the non-coding changes are cis-regulatory mutations causing a higher CDKN2A expression, whereas the missense mutations reduce the ability of ARF to interact with MDM2. Caspase assays for all genotypes revealed no apoptotic events and our results are consistent with a recent study indicating that the loss of melanocyte progenitors in Sex-linked barring in chicken is caused by premature differentiation and not apoptosis. Our results show that CDKN2A is a major locus driving the differentiation of avian melanocytes in a temporal and spatial manner.

Place, publisher, year, edition, pages
PUBLIC LIBRARY SCIENCE, 2017
Keywords
ARF TUMOR-SUPPRESSOR; GENE-EXPRESSION; LOCUS; MUTATIONS; INK4A/ARF; MELANOMA; GENOME; SENESCENCE; NICHE; CELLS
National Category
Genetics
Identifiers
urn:nbn:se:uu:diva-327066 (URN)10.1371/journal.pgen.1006665 (DOI)000402549200008 ()
Available from: 2017-08-01 Created: 2017-08-01 Last updated: 2018-01-03Bibliographically approved
Toto, A., Mattei, A., Jemth, P. & Gianni, S. (2017). Understanding the role of phosphorylation in the binding mechanism of a PDZ domain. Protein Engineering Design & Selection, 30(1), 1-5
Open this publication in new window or tab >>Understanding the role of phosphorylation in the binding mechanism of a PDZ domain
2017 (English)In: Protein Engineering Design & Selection, ISSN 1741-0126, E-ISSN 1741-0134, Vol. 30, no 1, p. 1-5Article in journal (Refereed) Published
Abstract [en]

The PDZ domain is one of the most common protein-protein interaction domains in mammalian species. While several studies have demonstrated the importance of phosphorylation in interactions involving PDZ domains, there is a paucity of detailed mechanistic data addressing how the PDZ interaction is affected by phosphorylation. Here, we address this question by equilibrium and kinetic binding experiments using PDZ2 from protein tyrosine phosphatase L1 and its interaction with a peptide from the natural ligand RIL. The results show that phosphorylation of a serine residue in the RIL peptide has dual and opposing effects: it increases both the association and dissociation rate constants, which leads to an overall weakening of binding. Furthermore, we performed binding experiments with a RIL peptide in which the serine was replaced by a glutamate, a commonly used method to mimic phosphorylation in proteins. Strikingly, both the affinity and the ionic strength dependence of the affinity differed markedly for the phosphoserine and glutamate peptides. These results show that, in this particular case, glutamate is a poor mimic of serine phosphorylation.

Place, publisher, year, edition, pages
OXFORD UNIV PRESS, 2017
Keywords
ionic strength, kinetics, peptide binding, protein-protein interaction
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:uu:diva-319870 (URN)10.1093/protein/gzw055 (DOI)000397129400001 ()27760803 (PubMedID)
Funder
Swedish Research Council
Available from: 2017-04-13 Created: 2017-04-13 Last updated: 2018-01-13Bibliographically approved
Gianni, S., Dogan, J. & Jemth, P. (2016). Coupled binding and folding of intrinsically disordered proteins: what can we learn from kinetics?. Current opinion in structural biology, 36, 18-24
Open this publication in new window or tab >>Coupled binding and folding of intrinsically disordered proteins: what can we learn from kinetics?
2016 (English)In: Current opinion in structural biology, ISSN 0959-440X, E-ISSN 1879-033X, Vol. 36, p. 18-24Article in journal (Refereed) Published
Abstract [en]

Protein or protein regions that are not forming well-defined structures in their free states under native-like conditions are called intrinsically disordered proteins. Such proteins are very common in protein-protein interactions, where their disorder apparently gives several advantages including optimal binding properties. To fully appreciate why protein disorder is advantageous for protein-protein interactions we need to understand the mechanism(s) of interaction. However, elucidating mechanisms in protein-protein interactions is usually very challenging. Here we discuss how kinetics in combination with protein engineering and structural information can be used to depict details of protein-protein interactions involving intrinsically disordered 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-294591 (URN)10.1016/j.sbi.2015.11.012 (DOI)000372681200005 ()26720267 (PubMedID)
Funder
Swedish Research Council, 2012-5096, 2014-4299
Available from: 2016-05-26 Created: 2016-05-25 Last updated: 2017-11-30Bibliographically 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
Gianni, S. & Jemth, P. (2016). Protein folding: Vexing debates on a fundamental problem. Biophysical Chemistry, 212, 17-21
Open this publication in new window or tab >>Protein folding: Vexing debates on a fundamental problem
2016 (English)In: Biophysical Chemistry, ISSN 0301-4622, E-ISSN 1873-4200, Vol. 212, p. 17-21Article, review/survey (Refereed) Published
Abstract [en]

The folding of proteins has been at the heart of protein chemistry and biophysics ever since the pioneering experiments by the labs of Fred Richards and Christian Anfinsen. But, despite nearly 60 years of intense research, there are unresolved issues and a lively debate regarding some aspects of this fundamental problem. In this review we give a personal account on some key topics in the field: (i) the nature of the denatured state of a protein, (ii) nucleation sites in the folding reaction, and (iii) the time it takes for individual molecules to traverse the transition state.

Keywords
Protein folding, Kinetics, Mutagenesis, Denatured state, Transition state, Transition path time
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-299179 (URN)10.1016/j.bpc.2016.03.001 (DOI)000374625600003 ()27018826 (PubMedID)
Funder
Swedish Research Council, 2012- 5096
Available from: 2016-07-15 Created: 2016-07-15 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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