Recognition of the DNA helix stabilizing anthramycin-N2 guanine adduct by UVRABC nuclease.
The binding of the anti-tumor antibiotic anthramycin to a defined linear DNA fragment was investigated using both exonuclease III and lambda exonuclease. We show that most of the guanine residues are reactive toward anthramycin; however, several guanine residues showed preferential reactivity for the drug. Using purified UVRA, UVRB and UVRC proteins we present evidence that these three proteins in concert are able to recognize and produce specific strand cleavage flanking anthramycin-DNA adducts. The cleavage of anthramycin adducts by UVRABC nuclease is specific and results in strand breaks at five or six bases 5' and three or four bases 3'-flanking an adduct. At some guanine residues single incisions were observed only on one side of the adduct. The 5' strand breaks observed often occurred as doublet bands on sequencing gels, indicating plasticity in the site of 5' cleavage whereas the 3' cleavage did not show this effect. When DNA fragments modified with elevated levels of anthramycin were used as substrates the activity of the UVRABC nuclease toward the anthramycin adducts decreased. Possible mechanisms for the recognition and specific cleavage of the helix-stabilizing anthramycin DNA adduct and other helix destabilizing lesions by the UVRABC nuclease are discussed.
Walter
R B
RB
University of Texas System Cancer Center, Science Park-Research Division, Smithville 78957.
Pierce
J
J
Case
R
R
Tang
M S
MS
D000876:Anthramycin / Q000378:metabolism*; D001483:Base Sequence; D001570:Benzodiazepinones / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics*; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D029968:Escherichia coli Proteins; D005090:Exodeoxyribonucleases / Q000378:metabolism; D006147:Guanine / Q000378:metabolism*; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010957:Plasmids; D014764:Viral Proteins
D001570:Benzodiazepinones; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D014764:Viral Proteins; D000876:Anthramycin; D006147:Guanine; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III; C043586:exo protein, Bacteriophage lambda; C036385:endodeoxyribonuclease uvrABC