Identification and characterization of the fourth single-stranded-DNA binding domain of replication protein A

Steven Brill, Suzanne Bastin-Shanower

Research output: Contribution to journalArticle

74 Citations (Scopus)

Abstract

Replication protein A (RPA), the heterotrimeric single-stranded-DNA (ssDNA) binding protein (SSB) of eukaryotes, contains two homologous ssDNA binding domains (A and B) in its largest subunit, RPA1, and a third domain in its second-largest subunit, RPA2. Here we report that Saccharomyces cerevisiae RPA1 contains a previously undetected ssDNA binding domain (domain C) lying in tandem with domains A and B. The carboxyterminal portion of domain C shows sequence similarity to domains A and B and to the region of RPA2 that binds ssDNA (domain D). The aromatic residues in domains A and B that are known to stack with the ssDNA bases are conserved in domain C, and as in domain A, one of these is required for viability in yeast. Interestingly, the amino-terminal portion of domain C contains a putative Cys4-type zinc-binding motif similar to that of another prokaryotic SSB, T4 gp32. We demonstrate that the ssDNA binding activity of domain C is uniquely sensitive to cysteine modification but that, as with gp32, ssDNA binding is not strictly dependent on zinc. The RPA heterotrimer is thus composed of at least four ssDNA binding domains and exhibits features of both bacterial and phage SSBs.

Original languageEnglish (US)
Pages (from-to)7225-7234
Number of pages10
JournalMolecular and cellular biology
Volume18
Issue number12
DOIs
StatePublished - Jan 1 1998

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Replication Protein A
Single-Stranded DNA
Zinc
DNA-Binding Proteins
Eukaryota
Bacteriophages
Cysteine
Saccharomyces cerevisiae
Carrier Proteins
Yeasts

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

Cite this

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abstract = "Replication protein A (RPA), the heterotrimeric single-stranded-DNA (ssDNA) binding protein (SSB) of eukaryotes, contains two homologous ssDNA binding domains (A and B) in its largest subunit, RPA1, and a third domain in its second-largest subunit, RPA2. Here we report that Saccharomyces cerevisiae RPA1 contains a previously undetected ssDNA binding domain (domain C) lying in tandem with domains A and B. The carboxyterminal portion of domain C shows sequence similarity to domains A and B and to the region of RPA2 that binds ssDNA (domain D). The aromatic residues in domains A and B that are known to stack with the ssDNA bases are conserved in domain C, and as in domain A, one of these is required for viability in yeast. Interestingly, the amino-terminal portion of domain C contains a putative Cys4-type zinc-binding motif similar to that of another prokaryotic SSB, T4 gp32. We demonstrate that the ssDNA binding activity of domain C is uniquely sensitive to cysteine modification but that, as with gp32, ssDNA binding is not strictly dependent on zinc. The RPA heterotrimer is thus composed of at least four ssDNA binding domains and exhibits features of both bacterial and phage SSBs.",
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Identification and characterization of the fourth single-stranded-DNA binding domain of replication protein A. / Brill, Steven; Bastin-Shanower, Suzanne.

In: Molecular and cellular biology, Vol. 18, No. 12, 01.01.1998, p. 7225-7234.

Research output: Contribution to journalArticle

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