METTL7A and METTL7B are responsible for S-thiol methyl transferase activity in liver

Abstract

S-Methylation of drugs containing thiol-moieties often alters their activity and results in detoxification. Historically, scientists attributed methylation of exogenous aliphatic and phenolic thiols to a putative S-adenosyl-L-methionine dependent membrane-associated phase II enzyme known as thiol methyltransferase (TMT). TMT has a broad substrate specificity and methylates the thiol metabolite of spironolactone, mertansine, ziprasidone, captopril, and the active metabolites of the thienopyridine pro-drugs, clopidogrel, and prasugrel. Despite TMTs role in the S-methylation of clinically relevant drugs, the enzyme(s) responsible for this activity remained unknown. We recently identified methyltransferase-like protein 7B (METTL7B) as an alkyl thiol-methyltransferase. METTL7B is an endoplasmic-reticulum-associated protein with similar biochemical properties and substrate specificity to TMT. Yet, the historic TMT inhibitor, 2,3-dichloro--methylbenzylamine (DCMB), has no effect on the activity of METTL7B, indicating that multiple enzymes contribute to TMT activity. Here we report that methyltransferase-like protein 7A (METTL7A), an uncharacterized member of the METTL7 family, also acts as a thiolmethyltransferase. METTL7A exhibits similar biochemical properties to TMT, including inhibition by DCMB (IC50 1.2 {micro}M). Applying quantitative proteomics to human liver microsomes and gene modulation experiments in HepG2 and HeLa cells, we determined that TMT activity correlates closely with METTL7A and METTL7B protein levels. Furthermore, purification of a novel His-GST-tagged recombinant protein and subsequent activity experiments prove that METTL7A can selectively methylate exogenous thiol-containing substrates, including 7-thiospironolactone, dithiothreitol, 4-chlorothiophenol, and mertansine. We conclude that the METTL7 family encodes for two enzymes, METTL7A and METTL7B, which we have renamed TMT1A1 and TMT1B1, respectively, that are responsible for TMT activity in liver microsomes. Significance StatementWe identified METTL7A (TMT1A1) and METTL7B (TMT1B1) as the enzymes responsible for the microsomal alkyl thiol methyltransferase activity. These are the first two enzymes directly associated with microsomal TMT activity. S-Methylation of many commonly prescribed thiol-containing drugs alters their pharmacological activity and/or toxicity, and identifying the enzymes responsible, will improve our understanding of the DMPK properties of alkyl- or phenolic-thiol-containing therapeutics. Visual Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=133 SRC="FIGDIR/small/511968v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@2419f9org.highwire.dtl.DTLVardef@254c5borg.highwire.dtl.DTLVardef@b81378org.highwire.dtl.DTLVardef@1064c79_HPS_FORMAT_FIGEXP M_FIG C_FIG