RT Journal Article SR Electronic T1 Tetrameric UvrD helicase is located at the E. coli replisome due to frequent replication blocks JF bioRxiv FD Cold Spring Harbor Laboratory SP 2021.02.22.432310 DO 10.1101/2021.02.22.432310 A1 Adam J. M Wollman A1 Aisha H. Syeda A1 Andrew Leech A1 Colin Guy A1 Peter McGlynn A1 Michelle Hawkins A1 Mark C. Leake YR 2021 UL http://biorxiv.org/content/early/2021/02/22/2021.02.22.432310.abstract AB DNA replication in all organisms must overcome nucleoprotein blocks to complete genome duplication. Accessory replicative helicases in Escherichia coli, Rep and UvrD, help replication machinery overcome blocks by removing incoming nucleoprotein complexes or aiding the re-initiation of replication. Mechanistic details of Rep function have emerged from recent live cell studies, however, the activities of UvrD in vivo remain unclear. Here, by integrating biochemical analysis and super-resolved single-molecule fluorescence microscopy, we discovered that UvrD self-associates into a tetramer and, unlike Rep, is not recruited to a specific replisome protein despite being found at approximately 80% of replication forks. By deleting rep and DNA repair factors mutS and uvrA, perturbing transcription by mutating RNA polymerase, and antibiotic inhibition; we show that the presence of UvrD at the fork is dependent on its activity. This is likely mediated by the very high frequency of replication blocks due to DNA bound proteins, including RNA polymerase, and DNA damage. UvrD is recruited to sites of nucleoprotein blocks via distinctly different mechanisms to Rep and therefore plays a more important and complementary role than previously realised in ensuring successful DNA replication.Competing Interest StatementThe authors have declared no competing interest.