title: Two DNA glycosylases in Escherichia coli which release primarily 3-methyladenine.
abstract: Two enzymes have been partially purified from Escherichia coli and designated 3-methyladenine DNA glycosylases I and II. The glycosylase I is that described by Riazuddin & Lindahl [Riazuddin, S., & Lindahl, T. (1978) Biochemistry 17, 2110-2118]. The apparent molecular weight of glycosylase I is 20 000, and that of II is 27 000. Glycosylase I releases 3-methyladenine (3-MeA) while II releases 3-MeA, 3-methylguanine (3-MeG), 7-methylguanine (7-MeG), and 7-methyladenine (7-MeA). The rate of release of 3-MeA by glycosylase II is 30 times that of 7-MeG. Glycosylase I is missing in mutants tag 1 and tag 2 [Karran, P., Lindahl, T., Ofsteng, I., Evenson, G. B., & Seeberg, E. (1980) J. Mol. Biol. 140, 101-127]. In crude extracts, the 3-MeA activity of II is approximately 10% of the total 3-MeA activity. A 50% inactivation at 48 degrees C required 5 min for I and 65 min for II. The apparent Km for 3-MeA residues for glycosylase I was 1.4 x 10(-8) M. The enzyme was inhibited noncompetitively by 3-MeA with an average apparent Ki of 1.6 mM. The apparent Km for 3-MeA, for glycosylase II, was 9.2 x 10(-9)M, and it was not inhibited by 3-MeA. The 3-MeA and 7-MeG activities of the glycosylase II preparation could not be separated by isoelectric focusing, by chromatography on DEAE, Sephadex G-100, phosphocellulose, DNA-cellulose, or carboxymethylcellulose, or by heating at 50 degrees C. The apparent Km for 7-MeG was 1.1 x 10(-8)M. Glycosylase II released N1-(carboxyethyl)adenine and N7-(carboxymethyl)guanine from DNA treated with beta-[3H]propiolactone but did not release the aflatoxin B-1 adduct at N-7 of guanine.
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000378:metabolism; D001426:Bacterial Proteins / Q000302:isolation & purification*; D045647:DNA Glycosylases; D004926:Escherichia coli / Q000201:enzymology*; D006147:Guanine / Q000031:analogs & derivatives / Q000378:metabolism; D006358:Hot Temperature; D007525:Isoelectric Focusing; D007700:Kinetics; D008970:Molecular Weight; D009699:N-Glycosyl Hydrolases / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; C025946:3-methyladenine; D006147:Guanine; C008450:7-methylguanine; C021565:3-methyladenine-DNA glycosylase; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C076131:DNA-3-methyladenine glycosidase II; D000225:Adenine