umu.sePublications
Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Arabidopsis thaliana peroxiredoxin Q is extraordinarily dynamic on the μs-ms timescale
Umeå University, Faculty of Science and Technology, Department of Chemistry. (Magnus Wolf-Watz)
Umeå University, Faculty of Science and Technology, Department of Chemistry. (Gerhard Gröbner)
Umeå University, Faculty of Science and Technology, Department of Chemistry. (Magnus Wolf-Watz)
Show others and affiliations
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Peroxiredoxin Q (PrxQ) isolated from Arabidopsis thaliana belongs to a family of redox enzymes called peroxiredoxins, which are thioredoxin- or glutaredoxin dependent peroxidases acting to reduce peroxides and in particular hydrogen peroxide. PrxQ cycles between an active reduced state and an inactive oxidized state during its catalytic cycle. The catalytic mechanism involves a nucleophilic attack of the catalytic cysteine on hydrogen peroxide to generate a sulfonic acid intermediate with a concerted release of a water molecule. This intermediate is subsequently relaxed by the reaction of a second cysteine, denoted as the resolving cysteine, generating an intermolecular disulphide bond to expel a second water molecule into solution. PrxQ is finally recycled to the active state by a thioredoxin dependent reduction. Previous structural studies of PrxQ homologues have provided the structural basis for the switch between reduced and oxidized conformations. Here we have performed a detailed study of the structure and dynamics of PrxQ in both the oxidized and reduced state. Reliable and experimentally validated structural models of PrxQ in both oxidation states were generated using homology based modeling. Model-free analyses of NMR spin relaxation show that PrxQ is monomeric in both oxidation states. As evident from fast R2 relaxation rates the reduced form of PrxQ undergoes unprecedented dynamics on the slow μs-ms timescale. The ground state of the conformational dynamics is likely the stably folded reduced state as implied by circular dichroism spectroscopy. We speculate that the extensive dynamics is intimately related to the catalytic function of PrxQ.

Keyword [en]
activity, circular dichroism, dynamics, enzyme, NMR
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Biophysics
Research subject
Biochemistry; Physical Chemistry
Identifiers
URN: urn:nbn:se:umu:diva-36751OAI: oai:DiVA.org:umu-36751DiVA: diva2:356108
Available from: 2010-10-11 Created: 2010-10-11 Last updated: 2010-10-15Bibliographically approved
In thesis
1. NMR studies of protein dynamics and structure
Open this publication in new window or tab >>NMR studies of protein dynamics and structure
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Enzymes are extraordinary molecules that can accelerate chemical reactions by several orders of magnitude. With recent advancements in structural biology together with classical enzymology the mechanism of many enzymes has become understood at the molecular level. During the last ten years significant efforts have been invested to understand the structure and dynamics of the actual catalyst (i. e. the enzyme). There has been a tremendous development in NMR spectroscopy (both hardware and pulse programs) that have enabled detailed studies of protein dynamics. In many cases there exists a strong coupling between enzyme dynamics and function. Here I have studied the conformational dynamics and thermodynamics of three model systems: adenylate kinase (Adk), Peroxiredoxin Q (PrxQ) and the structural protein S16. By developing a novel chemical shift-based method we show that Adk binds its two substrates AMP and ATP with an extraordinarily dynamic mechanism. For both substrate-saturated states the nucleotide-binding subdomains exchange between open and closed states, with the populations of these states being approximately equal. This finding contrasts with the traditional view of enzyme-substrate complexes as static low entropy states. We are also able to show that the individual subdomains in Adk fold and unfold in a non-cooperative manner. This finding is relevant from a functional perspective, since it allows a change in hydrogen bonding pattern upon substrate-binding without provoking global unfolding of the entire enzyme (as would be expected from a two-state folding mechanism). We also studied the structure and dynamics of the plant enzyme PrxQ in both reduced and oxidized states. Experimentally validated structural models were generated for both oxidation states. The reduced state displays unprecedented μs-ms conformational dynamics and we propose that this dynamics reflects local and functional unfolding of an α-helix in the active site. Finally, we solved the structure of S16 from Aquifex aeolicus and propose a model suggesting a link between thermostability and structure for a mesophilic and hyperthermophilic protein pair. A connection between the increased thermostability in the thermophilic S16 and residual structure in its unfolded state was discovered, persistent at high denaturant concentrations, thereby affecting the difference in heat capacity difference between the folded and unfolded state. In summary, we have contributed to the understanding of protein dynamics and to the coupling between dynamics and catalytic activity in enzymes.

Place, publisher, year, edition, pages
Umeå: Umeå universitet. Kemiska institutionen, 2010. 53 p.
Keyword
NMR, protein dynamics, relaxation, protein folding
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Physical Chemistry
Research subject
Biochemistry; Physical Chemistry
Identifiers
urn:nbn:se:umu:diva-36790 (URN)978-91-7459-092-0 (ISBN)
Public defence
2010-11-05, KBC-huset, KB3A9, 901 87 Umeå, Umeå, 10:00 (English)
Opponent
Supervisors
Available from: 2010-10-15 Created: 2010-10-11 Last updated: 2010-10-15Bibliographically approved

Open Access in DiVA

No full text

Authority records BETA

Ådén, JörgenWallgren, MarcusStorm, PatrikWeise, ChristophChristiansen, AlexanderSchröder, WolfgangFunk, ChristianeWolf-Watz, Magnus

Search in DiVA

By author/editor
Ådén, JörgenWallgren, MarcusStorm, PatrikWeise, ChristophChristiansen, AlexanderSchröder, WolfgangFunk, ChristianeWolf-Watz, Magnus
By organisation
Department of Chemistry
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)Biophysics

Search outside of DiVA

GoogleGoogle Scholar

urn-nbn

Altmetric score

urn-nbn
Total: 364 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf