Characterization of a cDNA encoding the 70-kDa single-stranded DNA-binding subunit of human replication protein A and the role of the protein in DNA replication

L. F. Erdile, W. D. Heyer, R. Kolodner, T. J. Kelly

Research output: Contribution to journalArticle

105 Scopus citations

Abstract

Replication protein A (RP-A) is a three-subunit single-stranded DNA-binding protein that has been isolated from human cells. RP-A is essential for SV40 DNA replication and may also be important in genetic recombination. The sequence of a cDNA encoding the 70-kDa subunit of human RP-A is reported. The 616-amino acid predicted open reading frame of the human protein is 31% identical with the 621-amino acid open reading frame of the 70-kDa subunit of RP-A from the yeast Saccharomyces cerevisiae. Both proteins share a highly conserved putative metal binding domain of the 4-cysteine type. The human cDNA directs production in Escherichia coli of a 70-kDa protein that reacts with a monoclonal antibody directed against the 70-kDa subunit of human RP-A. The recombinant 70-kDa subunit, purified from bacteria, exhibits single-stranded DNA binding activity comparable to that of the complete RP-A complex. The 70-kDa subunit is able to substitute for the complete human RP-A complex in stimulating the activity of DNA polymerase α-primase on a poly(dA)·oligo(dT) template. However, the 70-kDa subunit alone cannot substitute for the complete RP-A complex in SV40 DNA replication in vitro, suggesting an important functional role for the other subunits.

Original languageEnglish (US)
Pages (from-to)12090-12098
Number of pages9
JournalJournal of Biological Chemistry
Volume266
Issue number18
StatePublished - 1991
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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