title: Properties of the PriA helicase domain and its role in binding PriA to specific DNA structures.
abstract: Primosome assembly protein PriA functions in the assembly of the replisome at forked DNA structures. Whereas its N-terminal DNA binding domain (DBD) binds independently to DNA, the affinity of DBD protein for forked structures is relatively weak. Although the PriA helicase domain (HD) is required for high affinity fork binding, HD protein had very low affinity for DNA. It had only low levels of ATPase activity, and it hydrolyzed ATP when DNA was absent whereas PriA did not. HD catalyzed unwinding of a minimal substrate composed of a duplex with a 3' single-stranded tail. Single-strand binding protein (SSB) bound to the tail of this substrate inhibited this reaction by full-length PriA but enhanced the reaction by HD. SSB stabilized binding of PriA but not of DBD or HD to duplexes with a 5' or 3' single-stranded tail. On forked substrates SSB enhanced helicase action on the lagging-strand arm by PriA but not by HD. The results indicate that synergy of the DBD and HD allows stable binding at the interface between duplex and single-stranded DNA bound by SSB. This mode of binding may be analogous to fork binding, which orients the helicase to act on the lagging-strand side of the fork.
authors.affiliation: Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, Washington, D. C. 20057, USA.
authors.forename: Hua-Wei
authors.initials: HW
authors.lastname: Chen
authors.forename: Stella H
authors.initials: SH
authors.lastname: North
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nakai
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry; D001665:Binding Sites; D002384:Catalysis; D004247:DNA / Q000737:chemistry*; D004265:DNA Helicases / Q000737:chemistry*; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D006868:Hydrolysis; D008957:Models, Genetic; D010957:Plasmids / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology
chemical_list: D029968:Escherichia coli Proteins; D000255:Adenosine Triphosphate; D004247:DNA; D014357:Trypsin; D000251:Adenosine Triphosphatases; C506743:priA protein, E coli; D004265:DNA Helicases