The folate‐binding module of Thermus thermophilus cobalamin‐dependent methionine synthase displays a distinct variation of the classical TIM barrel: a TIM barrel with a `twist'

Methyl transfer between methyltetrahydrofolate and corrinoid molecules is a key reaction in biology that is catalyzed by a number of enzymes in many prokaryotic and eukaryotic organisms. One classic example of such an enzyme is cobalamin‐dependent methionine synthase (MS). MS is a large modular protein that utilizes an S N 2‐type mechanism to catalyze the chemically challenging methyl transfer from the tertiary amine (N5) of methyltetrahydrofolate to homocysteine in order to form methionine. Despite over half a century of study, many questions remain about how folate‐dependent methyltransferases, and MS in particular, function. Here, the structure of the folate‐binding (Fol) domain of MS from Thermus thermophilus is reported in the presence and absence of methyltetrahydrofolate. It is found that the methyltetrahydrofolate‐binding environment is similar to those of previously described methyltransferases, highlighting the conserved role of this domain in binding, and perhaps activating, the methyltetrahydrofolate substrate. These structural studies further reveal a new distinct and uncharacterized topology in the C‐terminal region of MS Fol domains. Furthermore, it is found that in contrast to the canonical TIM‐barrel β 8 α 8 fold found in all other folate‐binding domains, MS Fol domains exhibit a unique β 8 α 7 fold. It is posited that these structural differences are important for MS function.