Specific recognition of a multiply phosphorylated motif in the DNA repair scaffold XRCC1 by the FHA domain of human PNK

Nucleic Acids Res. 2009 Apr;37(5):1701-12. doi: 10.1093/nar/gkn1086. Epub 2009 Jan 20.

Abstract

Short-patch repair of DNA single-strand breaks and gaps (SSB) is coordinated by XRCC1, a scaffold protein that recruits the DNA polymerase and DNA ligase required for filling and sealing the damaged strand. XRCC1 can also recruit end-processing enzymes, such as PNK (polynucleotide kinase 3'-phosphatase), Aprataxin and APLF (aprataxin/PNK-like factor), which ensure the availability of a free 3'-hydroxyl on one side of the gap, and a 5'-phosphate group on the other, for the polymerase and ligase reactions respectively. PNK binds to a phosphorylated segment of XRCC1 (between its two C-terminal BRCT domains) via its Forkhead-associated (FHA) domain. We show here, contrary to previous studies, that the FHA domain of PNK binds specifically, and with high affinity to a multiply phosphorylated motif in XRCC1 containing a pSer-pThr dipeptide, and forms a 2:1 PNK:XRCC1 complex. The high-resolution crystal structure of a PNK-FHA-XRCC1 phosphopeptide complex reveals the basis for this unusual bis-phosphopeptide recognition, which is probably a common feature of the known XRCC1-associating end-processing enzymes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Binding Sites
  • Calorimetry
  • DNA Repair
  • DNA Repair Enzymes / chemistry*
  • DNA Repair Enzymes / metabolism
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Humans
  • Models, Molecular
  • Phosphopeptides / chemistry
  • Phosphopeptides / metabolism
  • Phosphoric Monoester Hydrolases / chemistry*
  • Phosphoric Monoester Hydrolases / metabolism
  • Phosphorylation
  • Phosphotransferases (Alcohol Group Acceptor) / chemistry*
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Sequence Deletion
  • X-ray Repair Cross Complementing Protein 1

Substances

  • DNA-Binding Proteins
  • Phosphopeptides
  • X-ray Repair Cross Complementing Protein 1
  • XRCC1 protein, human
  • PNKP protein, human
  • Phosphotransferases (Alcohol Group Acceptor)
  • Phosphoric Monoester Hydrolases
  • DNA Repair Enzymes