A Primase-Induced Conformational Switch Controls the Stability of the Bacterial Replisome

Enrico Monachino, Slobodan Jergic, Jacob S. Lewis, Zhi-Qiang Xu, Allen T.Y. Lo, Valerie L. O’Shea, James M. Berger, Nicholas E. Dixon, Antoine M. van Oijen

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

Recent studies of bacterial DNA replication have led to a picture of the replisome as an entity that freely exchanges DNA polymerases and displays intermittent coupling between the helicase and polymerase(s). Challenging the textbook model of the polymerase holoenzyme acting as a stable complex coordinating the replisome, these observations suggest a role of the helicase as the central organizing hub. We show here that the molecular origin of this newly found plasticity lies in the 500-fold increase in strength of the interaction between the polymerase holoenzyme and the replicative helicase upon association of the primase with the replisome. By combining in vitro ensemble-averaged and single-molecule assays, we demonstrate that this conformational switch operates during replication and promotes recruitment of multiple holoenzymes at the fork. Our observations provide a molecular mechanism for polymerase exchange and offer a revised model for the replication reaction that emphasizes its stochasticity.
Original languageEnglish (US)
Pages (from-to)140-154.e7
Number of pages1539999861
JournalMolecular Cell
Volume79
Issue number1
DOIs
StatePublished - Jul 2020
Externally publishedYes

Bibliographical note

KAUST Repository Item: Exported on 2021-03-11
Acknowledged KAUST grant number(s): OSR-2015-CRG4-2644
Acknowledgements: We are indebted to Karl Duderstadt and Christiaan Punter for ImageJ plugins, Yao Wang for providing protein reagents, Lisanne Spenkelink for development of and assistance with the sm-FRAP assays, and Harshad Ghodke for fruitful discussions. This work was supported by the Australian Research Council (grants DP150100956 and DP180100858 to A.M.v.O. and N.E.D. and Australian Laureate Fellowship FL140100027 to A.M.v.O.), King Abdullah University of Science and Technology, Saudi Arabia (grant OSR-2015-CRG4-2644 to N.E.D. and A.M.v.O), Nederlandse Organisatie voor Wetenschappelijk Onderzoek (grant 12CMCE03 to E.M.), and the National Institutes of Health (NIGMS grant R37-071747 to J.M.B.). Conceptualization, E.M. S.J. N.E.D. and A.M.v.O.; Methodology, E.M. S.J. J.S.L. N.E.D. and A.M.v.O.; Resources, E.M. S.J. J.S.L. Z.-Q.X. A.T.Y.L. and V.L.O.; Software, E.M.; Validation, Formal Analysis, & Writing – Original Draft, E.M. and S.J.; Investigation, E.M. S.J. and J.S.L.; Supervision, S.J. N.E.D. and A.M.v.O.; Writing – Review & Editing, E.M. S.J. J.M.B. N.E.D. and A.M.v.O.; Funding Acquisition, J.M.B. N.E.D. and A.M.v.O. The authors declare no competing interests.
This publication acknowledges KAUST support, but has no KAUST affiliated authors.

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology

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