title: Structures of human primase reveal design of nucleotide elongation site and mode of Pol α tethering.
abstract: Initiation of DNA synthesis in genomic duplication depends on primase, the DNA-dependent RNA polymerase that synthesizes de novo the oligonucleotides that prime DNA replication. Due to the discontinuous nature of DNA replication, primase activity on the lagging strand is required throughout the replication process. In eukaryotic cells, the presence of primase at the replication fork is secured by its physical association with DNA polymerase α (Pol α), which extends the RNA primer with deoxynucleotides. Our knowledge of the mechanism that primes DNA synthesis is very limited, as structural information for the eukaryotic enzyme has proved difficult to obtain. Here, we describe the crystal structure of human primase in heterodimeric form consisting of full-length catalytic subunit and a C-terminally truncated large subunit. We exploit the crystallographic model to define the architecture of its nucleotide elongation site and to show that the small subunit integrates primer initiation and elongation within the same set of functional residues. Furthermore, we define in atomic detail the mode of association of primase to Pol α, the critical interaction that keeps primase tethered to the eukaryotic replisome.
authors.affiliation: Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, United Kingdom.
authors.forename: Mairi Louise
authors.initials: ML
authors.lastname: Kilkenny
authors.forename: Michael Anthony
authors.initials: MA
authors.lastname: Longo
authors.forename: Rajika L
authors.initials: RL
authors.lastname: Perera
authors.forename: Luca
authors.initials: L
authors.lastname: Pellegrini
mesh_terms: D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004256:DNA Polymerase I / Q000737:chemistry* / Q000378:metabolism; D019915:DNA Primase / Q000737:chemistry* / Q000378:metabolism*; D004261:DNA Replication / Q000502:physiology*; D019281:Dimerization; D005454:Fluorescence Polarization; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation
chemical_list: D019915:DNA Primase; D004256:DNA Polymerase I
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