The architecture of an Okazaki fragment-processing holoenzyme from the archaeon Sulfolobus solfataricus

Giuseppe Cannone, Yuli Xu, Thomas R. Beattie, Stephen D. Bell, Laura Spagnolo*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

DNA replication on the lagging strand occurs via the synthesis and maturation of Okazaki fragments. In archaea and eukaryotes, the enzymatic activities required for this process are supplied by a replicative DNA polymerase, Flap endonuclease 1 (Fen1) and DNA ligase 1 (Lig1). These factors interact with the sliding clamp PCNA (proliferating cell nuclear antigen) providing a potential means of co-ordinating their sequential actions within a higher order assembly. In hyperthermophilic archaea of the Sulfolobus genus, PCNA is a defined heterotrimeric assembly and each subunit interacts preferentially with specific client proteins. We have exploited this inherent asymmetry to assemble a PCNA-polymerase-Fen1-ligase complex on DNA and have visualized it by electron microscopy. Our studies reveal the structural basis of co-occupancy of a single PCNA ring by the three distinct client proteins.

Original languageEnglish
Pages (from-to)239-245
Number of pages7
JournalBiochemical Journal
Volume465
DOIs
Publication statusPublished - 15 Jan 2015

Keywords

  • DNA Replication
  • Okazaki fragment maturation
  • Proliferating cell nuclear antigen (PCNA)
  • Single particle electron microscopy

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