title: Structure of a bifunctional DNA primase-polymerase.
abstract: Genome replication generally requires primases, which synthesize an initial oligonucleotide primer, and DNA polymerases, which elongate the primer. Primase and DNA polymerase activities are combined, however, in newly identified replicases from archaeal plasmids, such as pRN1 from Sulfolobus islandicus. Here we present a structure-function analysis of the pRN1 primase-polymerase (prim-pol) domain. The crystal structure shows a central depression lined by conserved residues. Mutations on one side of the depression reduce DNA affinity. On the opposite side of the depression cluster three acidic residues and a histidine, which are required for primase and DNA polymerase activity. One acidic residue binds a manganese ion, suggestive of a metal-dependent catalytic mechanism. The structure does not show any similarity to DNA polymerases, but is distantly related to archaeal and eukaryotic primases, with corresponding active-site residues. We propose that archaeal and eukaryotic primases and the prim-pol domain have a common evolutionary ancestor, a bifunctional replicase for small DNA genomes.
authors.affiliation: Institute of Biochemistry, University of Bayreuth, Universitätsstrasse 30, D-95447 Bayreuth, Germany. georg.lipps@uni-bayreuth.de
authors.forename: Georg
authors.initials: G
authors.lastname: Lipps
authors.forename: Andreas O
authors.initials: AO
authors.lastname: Weinzierl
authors.forename: Gudrun
authors.initials: G
authors.lastname: von Scheven
authors.forename: Claudia
authors.initials: C
authors.lastname: Buchen
authors.forename: Patrick
authors.initials: P
authors.lastname: Cramer
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D019915:DNA Primase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D016944:Sulfolobus / Q000201:enzymology
chemical_list: D017931:DNA Primers; D008670:Metals; D019915:DNA Primase; D004259:DNA-Directed DNA Polymerase