Human hepatic microsomal thiol methyltransferase: Assay conditions, biochemical properties, and correlation studies

Thiol methyltransferase (TMT) catalyzes the S-methylation of aliphatic sulfhydryl drugs and xenobiotic compounds. As a first step in the determination of whether genetic control of TMT activity in an easily accessible human cell, the red blood cell (RBC), might reflect the regulation of TMT in the human liver, we determined optimal conditions for the assay of human hepatic microsomal TMT activity. We then used those assay conditions to study the biochemical properties and regulation of TMT in the human liver for comparison with the properties of TMT in the human RBC. Substrate kinetic studies of hepatic microsomes performed with 2-mercaptoethanol (2-ME) revealed biphasic kinetics similar to those found in the human RBC, with apparent 'high-' and 'low'-affinity forms of TMT activity. The high-affinity form had an optimal pH of 7.2-7.6 and apparent K(M) values of 9.0 μM for 2- ME and 7.5 μM for S-adenosyl-L-methionine. The low-affinity form had an optimal pH of 8.8 and apparent K(M) values of 20 mM for 2-ME and 44 μM for S-adenosyl-L-methionine. Both forms were inhibited by compounds that also inhibited human RBC membrane TMT. The two kinetic forms of hepatic microsomal TMT activity were inactivated approximately 50% by heating for 15 min at 53°C and, as was found with RBC TMT, had very similar thermal stability profiles. Finally, as was also true of the RBC, there was a highly significant correlation between levels of activity of the two forms of TMT in 19 individual liver biopsy samples (r(s) = 0.961, p < 0.0001), but levels of activity of neither form of TMT were significantly correlated with those of a control microsomal enzyme, β-glucuronidase. Our results indicate that the biochemical properties of human hepatic microsomal TMT are very similar to those of human RBC membrane TMT.