Measurement of the removal of O6-methylguanine and O4-methylthymine from oligodeoxynucleotides using an immunoprecipitation technique

• Published in: Analytical biochemistry Vol. 183; no. 1; p. 74
• Main Authors: Mironov, N M, Wild, C P, Martel-Planche, G, Swann, P F, Montesano, R
• Format: Journal Article
• Language: English
• Published: United States 15-11-1989
• Subjects: Animals; Antibodies, Monoclonal; Base Sequence; Chromatography, High Pressure Liquid; Guanine - analogs & derivatives; Guanine - analysis; Humans; Male; Methyltransferases - metabolism; Molecular Sequence Data; O-Methylguanine-DNA Methyltransferase; Oligodeoxyribonucleotides - analysis; Precipitin Tests - methods; Rats; Thymine - analogs & derivatives; Thymine - analysis
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(89)90173-5

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.