Change search
ReferencesLink to record
Permanent link

Direct link
The Crystal Structure of Thermotoga maritima Class III Ribonucleotide Reductase Lacks a Radical Cysteine Pre-Positioned in the Active Site
Lund University.
Lund University.
Lund University.
Stockholm University. (Jarone Pinhassi)ORCID iD: 0000-0002-8779-6464
Show others and affiliations
2015 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 7, e0128199Article in journal (Refereed) Published
Abstract [en]

Ribonucleotide reductases (RNRs) catalyze the reduction of ribonucleotides to deoxyribonucleotides, the building blocks for DNA synthesis, and are found in all but a few organisms. RNRs use radical chemistry to catalyze the reduction reaction. Despite RNR having evolved several mechanisms for generation of different kinds of essential radicals across a large evolutionary time frame, this initial radical is normally always channelled to a strictly conserved cysteine residue directly adjacent to the substrate for initiation of substrate reduction, and this cysteine has been found in the structures of all RNRs solved to date. We present the crystal structure of an anaerobic RNR from the extreme thermophile Thermotoga maritima (tmNrdD), alone and in several complexes, including with the allosteric effector dATP and its cognate substrate CTP. In the crystal structure of the enzyme as purified, tmNrdD lacks a cysteine for radical transfer to the substrate pre-positioned in the active site. Nevertheless activity assays using anaerobic cell extracts from T. maritima demonstrate that the class III RNR is enzymatically active. Other genetic and microbiological evidence is summarized indicating that the enzyme is important for T. maritima. Mutation of either of two cysteine residues in a disordered loop far from the active site results in inactive enzyme. We discuss the possible mechanisms for radical initiation of substrate reduction given the collected evidence from the crystal structure, our activity assays and other published work. Taken together, the results suggest either that initiation of substrate reduction may involve unprecedented conformational changes in the enzyme to bring one of these cysteine residues to the expected position, or that alternative routes for initiation of the RNR reduction reaction may exist. Finally, we present a phylogenetic analysis showing that the structure of tmNrdD is representative of a new RNR subclass IIIh, present in all Thermotoga species plus a wider group of bacteria from the distantly related phyla Firmicutes, Bacteroidetes and Proteobacteria.

Place, publisher, year, edition, pages
2015. Vol. 10, no 7, e0128199
Keyword [en]
Sequence motif analysis, Crystal structure, Cysteine, Enzyme structure, Ribonucleotides, Chemical radicals, Electron density, Thermotoga
National Category
Structural Biology
Research subject
Natural Science
URN: urn:nbn:se:lnu:diva-50950DOI: 10.1371/journal.pone.0128199OAI: diva2:912986
Available from: 2016-03-18 Created: 2016-03-17 Last updated: 2016-04-08Bibliographically approved

Open Access in DiVA

fulltext(2579 kB)16 downloads
File information
File name FULLTEXT01.pdfFile size 2579 kBChecksum SHA-512
Type fulltextMimetype application/pdf

Other links

Publisher's full text

Search in DiVA

By author/editor
Lundin, Daniel
In the same journal
Structural Biology

Search outside of DiVA

GoogleGoogle Scholar
Total: 16 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 54 hits
ReferencesLink to record
Permanent link

Direct link