Roeland Boer, D.; Angel Ruiz-Maso, Jose; Rueda, Manuel; Petoukhov, Maxim V.; Machon, Cristina; Svergun, Dmitri I.; Orozco, Modesto; del Solar, Gloria; Coll, Miquel
Conformational plasticity of RepB, the replication initiator protein of promiscuous streptococcal plasmid pMV158
SCIENTIFIC REPORTS, 6 Art. No. 20915, FEB 15 2016

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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