title: Calf thymus DNA polymerase alpha-primase: "communication" and primer-template movement between the two active sites.
abstract: The DNA polymerase alpha-primase complex replicates single-stranded DNA by first synthesizing a short RNA primer (primase) which is then further elongated by the incorporation of dNTPs (DNA polymerase alpha). While primase and pol alpha function independently prior to synthesis of an RNA primer, the two activities become coordinated after primer synthesis. After primase generates a primer-template, it moves from the primase active site to the pol alpha active site for further elongation without dissociating into solution. Intramolecular transfer occurs immediately after primer synthesis and is employed on both long templates such as poly(dT) and short synthetic templates (< or = 60 nucleotides). Primer-template transfer and elongation by pol alpha are rapid compared to primer synthesis. After pol alpha elongates the primer, primase reinitiates primer synthesis, and the cycle is repeated. However, if dNTPs are absent such that primer elongation cannot occur, further primase activity is inhibited after a single round of primer synthesis. This "negative regulation" of primase activity is mediated by the newly generated primer-template provided the following conditions are met: (1) Primase synthesizes the primer; (2) the primer is 7-10 nucleotides long and remains bound to the template; (3) the template is of sufficient length; (4) the primer-template dissociates slowly from the enzyme complex; and (5) the primer-template interacts with the pol alpha active site. Polymerization of multiple dNTPs by pol alpha rapidly reactivates primase; hence, negative regulation of primase activity likely ensures a new primer is not synthesized until the previous one has been elongated by pol alpha.
authors.affiliation: Department of Chemistry and Biochemistry, University of Colorado, Boulder 80309-0215.
authors.forename: R J
authors.initials: RJ
authors.lastname: Sheaff
authors.forename: R D
authors.initials: RD
authors.lastname: Kuchta
authors.forename: D
authors.initials: D
authors.lastname: Ilsley
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D019915:DNA Primase; D004261:DNA Replication; D004789:Enzyme Activation; D008969:Molecular Sequence Data; D012316:RNA Nucleotidyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013698:Templates, Genetic; D013950:Thymus Gland / Q000201:enzymology*
chemical_list: D019915:DNA Primase; D012316:RNA Nucleotidyltransferases