Crystal structure of the Pyrococcus horikoshii DNA primase-UTP complex: implications for the mechanism of primer synthesis

Genes Cells. 2003 Dec;8(12):913-23. doi: 10.1111/j.1365-2443.2003.00693.x.

Abstract

Background: In chromosomal DNA replication, DNA primase initiates the synthesis of a dinucleotide on a single-stranded template DNA, and elongates it to form a primer RNA for the replicative DNA polymerase. Although the apo-structure of an archaeal primase has been reported, the mechanism of primer synthesis by the eukaryotic-type primase still remains to be elucidated.

Results: In this study, we present the crystal structure of the eukaryotic-type DNA primase from the hyperthermophilic archaeon (Pyrococcus horikoshii) with the uridine 5'-triphosphate (UTP). In the present primase-UTP complex, the primase binds the triphosphate moiety of the UTP at the active site, which includes Asp95, Asp97, and Asp280, the essential residues for the nucleotidyl transfer reaction.

Conclusion: The nucleotide binding geometry in this complex explains the previous biochemical analyses of the eukaryotic primase. Based on the complex structure, we constructed a model between the DNA primase and a primer/template DNA for the primer synthesis. This model facilitates the comprehension of the reported features of DNA primase.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Crystallography, X-Ray
  • DNA Damage
  • DNA Primase / chemistry*
  • DNA Primase / genetics
  • DNA Primase / metabolism
  • Deoxycytosine Nucleotides / chemistry
  • Deoxycytosine Nucleotides / metabolism
  • Macromolecular Substances
  • Models, Molecular*
  • Molecular Sequence Data
  • Mutation
  • Protein Binding
  • Pyrococcus horikoshii / enzymology*
  • RNA / biosynthesis
  • Sequence Alignment
  • Uridine Triphosphate / chemistry*
  • Uridine Triphosphate / metabolism

Substances

  • Deoxycytosine Nucleotides
  • Macromolecular Substances
  • RNA primers
  • RNA
  • DNA Primase
  • Uridine Triphosphate