Molecular cloning of the human gene, PNKP, encoding a polynucleotide kinase 3'-phosphatase and evidence for its role in repair of DNA strand breaks caused by oxidative damage.
Mammalian polynucleotide kinases catalyze the 5'-phosphorylation of nucleic acids and can have associated 3'-phosphatase activity, predictive of an important function in DNA repair following ionizing radiation or oxidative damage. The sequences of three tryptic peptides from a bovine 60-kDa polypeptide that correlated with 5'-DNA kinase and 3'-phosphatase activities identified human and murine dbEST clones. The 57.1-kDa conceptual translation product of this gene, polynucleotide kinase 3'-phosphatase (PNKP), contained a putative ATP binding site and a potential 3'-phosphatase domain with similarity to L-2-haloacid dehalogenases. BLAST searches identified possible homologs in Caenorhabditis elegans, Schizosaccharomyces pombe, and Drosophila melanogaster. The gene was localized to chromosome 19q13.3-13.4. Northern analysis indicated a 2-kilobase mRNA in eight human tissues. A glutathione S-transferase-PNKP fusion protein displayed 5'-DNA kinase and 3'-phosphatase activities. PNKP is the first gene for a DNA-specific kinase from any organism. PNKP expression partially rescued the sensitivity to oxidative damaging agents of the Escherichia coli DNA repair-deficient xth nfo double mutant. PNKP gene function restored termini suitable for DNA polymerase, consistent with in vivo removal of 3'-phosphate groups, facilitating DNA repair.
Jilani
A
A
Molecular Oncology Group, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, Montreal, Quebec H3T 1E2.
Ramotar
D
D
Slack
C
C
Ong
C
C
Yang
X M
XM
Scherer
S W
SW
Lasko
D D
DD
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002417:Cattle; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004249:DNA Damage; D004260:DNA Repair; D018076:DNA, Complementary / Q000737:chemistry; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics* / Q000502:physiology; D011116:Polynucleotide 5'-Hydroxyl-Kinase / Q000378:metabolism*
D018076:DNA, Complementary; D006861:Hydrogen Peroxide; D011116:Polynucleotide 5'-Hydroxyl-Kinase; D010744:Phosphoric Monoester Hydrolases