Structural basis for non-radical catalysis by TsrM, a radical SAM methylase

Crystal structures of a cobalamin-dependent radical S-adenosylmethionine (SAM) methylase reveal an unexpected mechanism that involves substrate-assisted catalysis whereby the carboxylate group of the co-substrate SAM serves as a general base. Tryptophan 2C methyltransferase (TsrM) methylates C2 of the indole ring of l-tryptophan during biosynthesis of the quinaldic acid moiety of thiostrepton. TsrM is annotated as a cobalamin-dependent radical S-adenosylmethionine (SAM) methylase; however, TsrM does not reductively cleave SAM to the universal 5MODIFIER LETTER PRIME-deoxyadenosyl 5MODIFIER LETTER PRIME-radical intermediate, a hallmark of radical SAM (RS) enzymes. Herein, we report structures of TsrM from Kitasatospora setae, which are the first structures of a cobalamin-dependent radical SAM methylase. Unexpectedly, the structures show an essential arginine residue that resides in the proximal coordination sphere of the cobalamin cofactor, and a [4Fe-4S] cluster that is ligated by a glutamyl residue and three cysteines in a canonical CXXXCXXC RS motif. Structures in the presence of substrates suggest a substrate-assisted mechanism of catalysis, wherein the carboxylate group of SAM serves as a general base to deprotonate N1 of the tryptophan substrate, facilitating the formation of a C2 carbanion.

Automated summary

The crystal structures of a cobalamin-dependent radical S-adenosylmethionine (SAM) methylase reveal an unexpected mechanism involving substrate-assisted catalysis with the carboxylate group of SAM serving as a general base. Tryptophan 2C methyltransferase (TsrM) methylates C2 of the indole ring of l-tryptophan during biosynthesis of thiostrepton's quinaldic acid moiety. The structures of TsrM from Kitasatospora setae show an essential arginine residue near the cobalamin cofactor, and a [4Fe-4S] cluster ligated by a glutamyl residue and three cysteines in a canonical CXXXCXXC RS motif. Substrate-assisted catalysis is suggested by structures in the presence of substrates, where the carboxylate group of SAM acts as a general base to deprotonate N1 of the tryptophan substrate, facilitating the formation of a C2 carbanion.