Bulk phase biochemistry of PIF1 and RecQ4 family helicases.

ABSTRACT

DNA helicases are involved in nearly all facets of genome integrity, and in humans, mutations in helicase-encoding genes are often linked to diseases of genomic instability. Two highly studied and evolutionarily conserved helicase families are the PIF1 and RecQ helicases. Enzymes in these families have known roles in DNA replication, recombination, and repair, as well as telomere maintenance, DNA recombination, and transcription. Although genetics, structural biology, and a variety of other techniques have been used to study these helicases, ensemble analyses of their basic biochemical activities such as DNA binding, ATP hydrolysis, and DNA unwinding have made significant contributions to our understanding of their physiological roles. Here, we present general methods to generate recombinant proteins from both helicase families, as well as standard biochemical assays to investigate their activities on DNA.