Role of zinc-finger motif in redox regulation of human replication protein A

M. Wang, J. S. You, S. H. Lee

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Abstract

Replication protein A (RPA) is a heterotrimeric zinc-finger protein complex involved in DNA replication, repair, and genetic recombination. Unlike other zinc-finger proteins, RPA's zinc-finger motif is not essential for its single-stranded DNA (ssDNA) binding activity, but is involved in redox regulation of its single-stranded DNA (ss-DNA) binding activity. To get an insight into the regulation of RPA-ssDNA interaction, wild-type RPA (wt-RPA) and zinc-finger mutant were examined for ssDNA binding activity using surface plasmon resonance technique. Interaction of wt-RPA with ssDNA under nonreducing conditions was very weak (KD × 2.33 × 10-8 M) compared with that under reducing conditions (KD = 7.35 × 10-11 M), whereas ssDNA binding affinity of the zinc-finger mutant was not affected by redox. The divalent ion chelator, o-phenanthroline, significantly reduced wt-RPA-ss-DNA interaction, but had no effect on the zinc-finger mutant. The inhibitory effect of o-phenanthroline on RPA-ss-DNA interaction was reversed by Zn(II), but not by other divalent cations, suggesting that Zn(II) is the unique metal coordinating the zinc-finger cysteines in redox regulation of RPA-ssDNA interaction. In DNA repair, redox affected RPA's interaction with damaged DNA, but not its role in stabilizing the xeroderma pigmentosum group A (XPA)-damaged DNA complex, suggesting that the zinc-finger motif may mediate the transition of RPA-XPA interaction to a stable RPA-XPA-damaged DNA complex in a redox-dependent manner.

Original languageEnglish (US)
Pages (from-to)657-669
Number of pages13
JournalAntioxidants and Redox Signaling
Volume3
Issue number4
DOIs
StatePublished - Jan 1 2001

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ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Molecular Biology
  • Clinical Biochemistry
  • Cell Biology

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