2 research outputs found
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The signature motif of the Saccharomyces cerevisiae Pif1 DNA helicase is essential in vivo for mitochondrial and nuclear functions and in vitro for ATPase activity
Pif1 family DNA helicases are conserved from bacteria to humans and have critical and diverse functions in vivo that promote genome integrity. Pif1 family helicases share a 23 amino acid region, called the Pif1 signature motif (SM) that is unique to this family. To determine the importance of the SM, we did mutational and functional analysis of the SM from the Saccharomyces cerevisiae Pif1 (ScPif1). The mutations deleted portions of the SM, made one or multiple single amino acid changes in the SM, replaced the SM with its counterpart from a bacterial Pif1 family helicase and substituted an α-helical domain from another helicase for the part of the SM that forms an α helix. Mutants were tested for maintenance of mitochondrial DNA, inhibition of telomerase at telomeres and double strand breaks, and promotion of Okazaki fragment maturation. Although certain single amino acid changes in the SM can be tolerated, the presence and sequence of the ScPif1 SM were essential for all tested in vivo functions. Consistent with the in vivo analyses, in vitro studies showed that the presence and sequence of the ScPif1 SM were critical for ATPase activity but not substrate binding
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Getting it done at the ends: Pif1 family DNA helicases and telomeres
It is widely appreciated that the ends of linear DNA molecules cannot be fully replicated by the
conventional replication apparatus. Less well known is that semi-conservative replication of
telomeric DNA also presents problems for DNA replication. These problems likely arise from the
atypical chromatin structure of telomeres, the GC-richness of telomeric DNA that makes it prone
to forming DNA secondary structures, and from RNA-DNA hybrids, formed by transcripts of one
or both DNA strands. Given the different aspects of telomeres that complicate their replication, it
is not surprising that multiple DNA helicases promote replication of telomeric DNA. This review
focuses on one such class of DNA helicases, the Pif1 family of 5′–3′ DNA helicases. In budding
and fission yeasts, Pif1 family helicases impact both telomerase-mediated and semi-conservative
replication of telomeric DNA as well as recombination-mediated telomere lengthening