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Nucleic Acids Res. 2007;35(17):5809-18. Epub 2007 Aug 24.

The yeast Pif1p DNA helicase preferentially unwinds RNA DNA substrates.

Author information

1
Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA. jboule@princeton.edu

Abstract

Pif1p is the prototypical member of the PIF1 family of DNA helicases, a subfamily of SFI helicases conserved from yeast to humans. Baker's yeast Pif1p is involved in the maintenance of mitochondrial, ribosomal and telomeric DNA and may also have a general role in chromosomal replication by affecting Okazaki fragment maturation. Here we investigate the substrate preferences for Pif1p. The enzyme was preferentially active on RNA-DNA hybrids, as seen by faster unwinding rates on RNA-DNA hybrids compared to DNA-DNA hybrids. When using forked substrates, which have been shown previously to stimulate the enzyme, Pif1p demonstrated a preference for RNA-DNA hybrids. This preferential unwinding could not be correlated to preferential binding of Pif1p to the substrates that were the most readily unwound. Although the addition of the single-strand DNA-binding protein replication protein A (RPA) stimulated the helicase reaction on all substrates, it did not diminish the preference of Pif1p for RNA-DNA substrates. Thus, forked RNA-DNA substrates are the favored substrates for Pif1p in vitro. We discuss these findings in terms of the known biological roles of the enzyme.

PMID:
17720711
PMCID:
PMC2034482
DOI:
10.1093/nar/gkm613
[Indexed for MEDLINE]
Free PMC Article

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