Measurement of the removal of O6-methylguanine and O4-methylthymine from oligodeoxynucleotides using an immunoprecipitation technique.
A sensitive and rapid procedure for measurement of alkyltransferase repair activity involving oligodeoxynucleotides followed by immunoprecipitation is described. Dodecadeoxynucleotides containing O6-methylguanine or O4-methylthymine were used as substrates for alkyltransferases and the reaction products of methylated or demethylated substrates were separated by precipitation with highly specific antibodies. This approach for O6-alkylguanine-DNA alkyltransferase measurement is far more rapid than when the reaction products are separated by chromatography. This technique makes the assay applicable to large-scale epidemiological or clinical studies and suggests a similar methodology could be applied for other DNA repair enzymes.
Mironov
N M
NM
Unit of Mechanisms of Carcinogenesis, International Agency for Research on Cancer, Lyon, France.
Wild
C P
CP
Martel-Planche
G
G
Swann
P F
PF
Montesano
R
R
D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D006147:Guanine / Q000031:analogs & derivatives* / Q000032:analysis; D006801:Humans; D008297:Male; D008780:Methyltransferases / Q000378:metabolism; D008969:Molecular Sequence Data; D019853:O(6)-Methylguanine-DNA Methyltransferase; D009838:Oligodeoxyribonucleotides / Q000032:analysis; D011233:Precipitin Tests / Q000379:methods; D051381:Rats; D013941:Thymine / Q000031:analogs & derivatives* / Q000032:analysis
D000911:Antibodies, Monoclonal; D009838:Oligodeoxyribonucleotides; C021223:O-4-methylthymine; D006147:Guanine; C008449:O-(6)-methylguanine; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D013941:Thymine