Reconstitution of simian virus 40 DNA replication with purified proteins

D. H. Weinberg, K. L. Collins, P. Simancek, A. Russo, M. S. Wold, D. M. Virshup, Thomas Kelly

Research output: Contribution to journalArticle

Abstract

Replication of plasmid DNA molecules containing the simian virus 40 (SV40) origin of DNA replication has been reconstituted with seven highly purified cellular proteins plus the SV40 large tumor (T) antigen. Initiation of DNA synthesis is absolutely dependent upon T antigen, replication protein A, and the DNA polymerase α-primase complex and is stimulated by the catalytic subunit of protein phosphatase 2A. Efficient elongation of nascent chains additionally requires proliferating cell nuclear antigen, replication factor C, DNA topoisomerase I, and DNA polymerase δ. Electron microscopic studies indicate that DNA replication begins at the viral origin and proceeds via intermediates containing two forks that move in opposite directions. These findings indicate that the reconstituted replication reaction has many of the characteristics expected of authentic viral DNA replication.

Original languageEnglish (US)
Pages (from-to)8692-8696
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume87
Issue number22
DOIs
Publication statusPublished - 1990
Externally publishedYes

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Keywords

  • DNA polymerase δ
  • eukaryotic DNA replication
  • replication factor C

ASJC Scopus subject areas

  • Genetics
  • General

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