Physicochemical studies of human O6‐methylguanine‐DNA methyltransferase

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Abstract
O 6‐Methylguanine‐DNA methyltransferase, present in most organisms, removes mutagenic and carcinogenic O6‐alkylguanine from DNA by accepting the alkyl group in a stoichiometric reaction. The protein has been partially purified from human placenta. It reacts with second‐order rate constants of 2.20 × 108 and 0.067 × 108 lmol‐1 min‐1 at 37°C for duplex and single‐stranded DNA substrates, respectively. The corresponding value for the alkylated base in synthetic poly(dC, dG, m6dG) is 0.02 × 108 lmol−1 min−1. The native protein is monomeric with a molecular mass of 22–24 kDa. Methylation of the protein does not lead to a gross change in its conformation but causes a slight reduction in its isoelectric point of 6.2. Although DNA protects the protein from heat inactivation, both duplex and single‐stranded DNAs inhibit its activity in a concentration‐dependent manner. The transferase reaction rate is also strongly inhibited by salt with about 20% of the maximum rate observed in physiological ionic strength. This inhibition is nonspecific with respect to the ions of univalent salts.

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