The DNA repair endonuclease XPG binds to proliferating cell nuclear antigen (PCNA) and shares sequence elements with the PCNA-binding regions of FEN-1 and cyclin-dependent kinase inhibitor p21.
Proliferating cell nuclear antigen (PCNA) is a DNA polymerase accessory factor that is required for DNA replication during S phase of the cell cycle and for resynthesis during nucleotide excision repair of damaged DNA. PCNA binds to flap endonuclease 1 (FEN-1), a structure-specific endonuclease involved in DNA replication. Here we report the direct physical interaction of PCNA with xeroderma pigmentosum (XP) G, a structure-specific repair endonuclease that is homologous to FEN-1. We have identified a 28-amino acid region of human FEN-1 (residues 328-355) and a 29-amino acid region of human XPG (residues 981-1009) that contains the PCNA binding activity. These regions share key hydrophobic residues with the PCNA-binding domain of the cyclin-dependent kinase inhibitor p21(Waf1/Cip1), and all three competed with one another for binding to PCNA. A conserved arginine in FEN-1 (Arg339) and XPG (Arg992) was found to be crucial for PCNA binding activity. R992A and R992E mutant forms of XPG failed to fully reconstitute nucleotide excision repair in an in vivo complementation assay. These results raise the possibility of a mechanistic linkage between excision and repair synthesis that is mediated by PCNA.
Gary
R
R
Life Sciences Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA.
Ludwig
D L
DL
Cornelius
H L
HL
MacInnes
M A
MA
Park
M S
MS
D000595:Amino Acid Sequence; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000737:chemistry / Q000378:metabolism*; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000378:metabolism*; D004720:Endonucleases; D045585:Flap Endonucleases; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009687:Nuclear Proteins; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors; D000099250:DNA Excision Repair Protein ERCC-5
D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004268:DNA-Binding Proteins; D004706:Endodeoxyribonucleases; D004720:Endonucleases; D045585:Flap Endonucleases; D009687:Nuclear Proteins; D018809:Proliferating Cell Nuclear Antigen; D014157:Transcription Factors; D000099250:DNA Excision Repair Protein ERCC-5; C493956:CDKN1A protein, human; C468317:FEN1 protein, human