RNA primer handoff in bacteriophage T4 DNA replication: The role of single-stranded DNA-binding protein and polymerase accessory proteins

Scott W. Nelson, Ravindra Kumar, Stephen J. Benkovic

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

In T4 phage, coordinated leading and lagging strand DNA synthesis is carried out by an eight-protein complex termed the replisome. The control of lagging strand DNA synthesis depends on a highly dynamic replisome with several proteins entering and leaving during DNA replication. Here we examine the role of single-stranded binding protein (gp32) in the repetitive cycles of lagging strand synthesis. Removal of the protein-interacting domain of gp32 results in a reduction in the number of primers synthesized and in the efficiency of primer transfer to the polymerase. We find that the primase protein is moderately processive, and this processivity depends on the presence of full-length gp32 at the replication fork. Surprisingly, we find that an increase in the efficiency of primer transfer to the clamp protein correlates with a decrease in the dissociation rate of the primase from the replisome. These findings result in a revised model of lagging strand DNA synthesis where the primase remains as part of the replisome after each successful cycle of Okazaki fragment synthesis. A delay in primer transfer results in an increased probability of the primase dissociating from the replication fork. The interplay between gp32, primase, clamp, and clamp loader dictates the rate and efficiency of primer synthesis, polymerase recycling, and primer transfer to the polymerase.

Original languageEnglish (US)
Pages (from-to)22838-22846
Number of pages9
JournalJournal of Biological Chemistry
Volume283
Issue number33
DOIs
StatePublished - Aug 15 2008

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DNA Primase
Bacteriophage T4
Bacteriophages
DNA-Binding Proteins
Accessories
DNA Replication
DNA
Clamping devices
Proteins
Recycling
Loaders
Carrier Proteins
RNA primers

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

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RNA primer handoff in bacteriophage T4 DNA replication : The role of single-stranded DNA-binding protein and polymerase accessory proteins. / Nelson, Scott W.; Kumar, Ravindra; Benkovic, Stephen J.

In: Journal of Biological Chemistry, Vol. 283, No. 33, 15.08.2008, p. 22838-22846.

Research output: Contribution to journalArticle

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