Properties of an endonuclease activity in Micrococcus luteus acting on gamma-irradiated DNA and on apurinic DNA.
A protein fraction from Micrococcus luteus with endonuclease activity against gamma-irradiated DNA was isolated and characterized. An additional activity on apurinic sites could not be separated, either by sucrose gradient sedimentation or by gel filtration through Sephadex G 100. From gel filtration, a molecular weight of about 25 000 was calculated for both endonuclease activities. The endonuclease activity against gamma-irradiated DNA was stimulated five-fold with 5 mM Mg++, whereas that against apurinic sites was less dependent on the Mg++ concentration. 100 mM KCl inhibited the gamma-ray endonuclease, but not the apurinic endonuclease activity. In gamma-irradiated RNA the protein recognized 1.65 endonuclease sensitive sites per radiation induced single-strand break, among which are 0.45 alkali labile lesions in the nucleotide strand. The affinity of the enzyme for the endonuclease sensitive site was evaluated resulting in a Km-value of 73 nM.
D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D004269:DNA, Bacterial / Q000378:metabolism / Q000528:radiation effects*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004307:Dose-Response Relationship, Radiation; D004720:Endonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D005720:Gamma Rays; D008274:Magnesium / Q000494:pharmacology; D008837:Micrococcus / Q000201:enzymology*; D011687:Purines; D014965:X-Rays
D004269:DNA, Bacterial; D011687:Purines; D004259:DNA-Directed DNA Polymerase; D004720:Endonucleases; D008274:Magnesium