Conformational plasticity of RepB, the replication initiator protein of promiscuous streptococcal plasmid pMV158

DNA replication initiation is a vital and tightly regulated step in all replicons and requires an initiator factor that specifically recognizes the DNA replication origin and starts replication. RepB from the promiscuous streptococcal plasmid pMV158 is a hexameric ring protein evolutionary related t...

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Bibliographic Details
Authors: Boer, Roeland, Ruiz Masó, José Angel, Petoukhov, Maxim, Machón Sobrado, Cristina, Svergun, Dmitri, Orozco López, Modesto, Solar, Gloria del, Coll, Miquel
Format: article
Status:Published version
Publication Date:2016
Country:España
Institution:Universidad de Barcelona
Repository:Dipòsit Digital de la UB
OAI Identifier:oai:diposit.ub.edu:2445/114049
Online Access:https://hdl.handle.net/2445/114049
Access Level:Open access
Keyword:Duplicació de l'ADN
Models moleculars
Radiocristal·lografia
DNA replication
Molecular models
X-ray crystallography
Description
Summary:DNA replication initiation is a vital and tightly regulated step in all replicons and requires an initiator factor that specifically recognizes the DNA replication origin and starts replication. RepB from the promiscuous streptococcal plasmid pMV158 is a hexameric ring protein evolutionary related to viral initiators. Here we explore the conformational plasticity of the RepB hexamer by i) SAXS, ii) sedimentation experiments, iii) molecular simulations and iv) X-ray crystallography. Combining these techniques, we derive an estimate of the conformational ensemble in solution showing that the C-terminal oligomerisation domains of the protein form a rigid cylindrical scaffold to which the N-terminal DNA-binding/catalytic domains are attached as highly flexible appendages, featuring multiple orientations. In addition, we show that the hinge region connecting both domains plays a pivotal role in the observed plasticity. Sequence comparisons and a literature survey show that this hinge region could exists in other initiators, suggesting that it is a common, crucial structural element for DNA binding and manipulation.

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