Completion of mammalian lagging strand DNA replication using purified proteins

J. J. Turchi, R. A. Bambara

Research output: Contribution to journalArticle

69 Scopus citations

Abstract

We have modeled the reactions involved in the completion of lagging strand DNA replication using a synthetic DNA substrate and purified enzymes from calf thymus. We have demonstrated that each polymerase, α, δ, and ε, is capable of extending an upstream 3' terminus to generate a nick in the DNA substrate that is subsequently ligated by DNA ligase I. Synthesis by each polymerase and subsequent ligation occurred efficiently after the addition of the 50-kDa 5'- to 3'-exonuclease. Analyses are presented which show that a substantial proportion of the ligated products is the result of polymerase ε, exonuclease, and DNA ligase I, all acting on the same DNA template. That is, polymerase fills in the gap, then both the polymerase and exonuclease act, one adding and the other removing nucleotides, followed by ligation. Results presented suggest that polymerase α, δ, or ε may functionally interact with DNA ligase I and the 5'- to 3'-exonuclease to perform the enzymatic reactions required for the completion of lagging strand DNA synthesis.

Original languageEnglish (US)
Pages (from-to)15136-15141
Number of pages6
JournalJournal of Biological Chemistry
Volume268
Issue number20
StatePublished - 1993

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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