Human nucleotide excision nuclease incises synthetic double-stranded DNA containing a pyrimidine dimer at the fourth phosphodiester linkage 3' to the pyrimidine dimer.
Linear 75mer double-stranded DNA containing a single pyrimidine dimer at a unique site was used to investigate pyrimidine dimer-dependent endonuclease activities from human cells. HeLaS3 cell extract incised the target DNA at the fourth phosphodiester linkage 3' to the pyrimidine dimer. However, incision of the DNA at 5' side of the pyrimidine dimer was not detected. The incision was also detected in cell extracts prepared from other excision repair-proficient cell lines. Incision was detected only on the DNA strand containing a pyrimidine dimer in the presence of poly(dI-dC)-poly(dI- dC) double strand. The reaction required Mg2+ but not ATP. The extract prepared from excision repair-deficient xeroderma pigmentosum (XP) cells belonging to the complementation group A was unable to incise the DNA. Extracts from the complementation groups C, D, and G incised the DNA very weakly at the third phosphodiester linkage 3' to the pyrimidine dimer, a site different from that incised by normal human cell extract. These results suggest that the observed incision reaction is associated with excision repair in human cells.
Tateishi
S
S
Institute of Molecular Embryology and Genetics, Kumamoto University School of Medicine, Japan.
Hori
N
N
Ohtsuka
E
E
Yamaizumi
M
M
D000255:Adenosine Triphosphate / Q000494:pharmacology; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000378:metabolism*; D004249:DNA Damage; D004260:DNA Repair; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis; D011089:Polydeoxyribonucleotides / Q000378:metabolism; D011740:Pyrimidine Dimers; D013379:Substrate Specificity; D014983:Xeroderma Pigmentosum / Q000201:enzymology
D029968:Escherichia coli Proteins; D009838:Oligodeoxyribonucleotides; D011089:Polydeoxyribonucleotides; D011740:Pyrimidine Dimers; C031156:poly d(I-C); D000255:Adenosine Triphosphate; D004247:DNA; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; D008274:Magnesium