title: Human DNA polymerase α in binary complex with a DNA:DNA template-primer.
abstract: The Polα/primase complex assembles the short RNA-DNA fragments for priming of lagging and leading strand DNA replication in eukaryotes. As such, the Polα polymerase subunit encounters two types of substrates during primer synthesis: an RNA:DNA helix and a DNA:DNA helix. The engagement of the polymerase subunit with the DNA:DNA helix has been suggested as the of basis for primer termination in eukaryotes. However, there is no structural information on how the Polα polymerase subunit actually engages with a DNA:DNA helix during primer synthesis. We present here the first crystal structure of human Polα polymerase subunit in complex with a DNA:DNA helix. Unexpectedly, we find that portion of the DNA:DNA helix in contact with the polymerase is not in a B-form but in a hybrid A-B form. Almost all of the contacts observed previously with an RNA primer are preserved with a DNA primer--with the same set of polymerase residues tracking the sugar-phosphate backbone of the DNA or RNA primer. Thus, rather than loss of specific contacts, the free energy cost of distorting DNA from B- to hybrid A-B form may augur the termination of primer synthesis in eukaryotes.
authors.affiliation: Department of Structural &Chemical Biology, Mount Sinai School of Medicine, Box 1677, 1425 Madison Avenue, New York, NY 10029, USA.
authors.forename: Javier
authors.initials: J
authors.lastname: Coloma
authors.affiliation: Department of Biochemistry and Molecular Biology, 301 University Boulevard, University of Texas Medical Branch, Galveston, TX 77755-1061, USA.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Johnson
authors.affiliation: Department of Biochemistry and Molecular Biology, 301 University Boulevard, University of Texas Medical Branch, Galveston, TX 77755-1061, USA.
authors.forename: Louise
authors.initials: L
authors.lastname: Prakash
authors.affiliation: Department of Biochemistry and Molecular Biology, 301 University Boulevard, University of Texas Medical Branch, Galveston, TX 77755-1061, USA.
authors.forename: Satya
authors.initials: S
authors.lastname: Prakash
authors.affiliation: Department of Structural &Chemical Biology, Mount Sinai School of Medicine, Box 1677, 1425 Madison Avenue, New York, NY 10029, USA.
authors.forename: Aneel K
authors.initials: AK
authors.lastname: Aggarwal
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004256:DNA Polymerase I / Q000737:chemistry / Q000378:metabolism*; D017931:DNA Primers; D004261:DNA Replication; D005454:Fluorescence Polarization; D006801:Humans; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation; D000072417:Protein Domains; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013698:Templates, Genetic
chemical_list: D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D004247:DNA; D004256:DNA Polymerase I