Fig. 3: Structural insight into the mechanism of biphytanyl chain formation.
From: Discovery, structure and mechanism of a tetraether lipid synthase
a, The active site of GDGT–MAS binds two archaeal lipids, L1P (yellow) and AG (orange), and directs one carbon chain from each lipid into separate pockets. The pocket that leads to 5′-dAH and the [Fe4S4]C cluster is the proposed reaction centre of GDGT–MAS. b, GDGT–MAS catalyses the formation of the biphytanyl chain by coupling two terminal Csp3 carbons, which our substrate-bound structure indicates are 9.9 Å apart. Generation of the Csp3–Csp3 bond would necessitate two sequential C–H activations and storage of a high-energy substrate radical intermediate. The substrate-bound complex suggests two potential mechanisms for storage of the high-energy radical intermediate: formation of (1) a terminal olefin and (2) an [Fe4S4]C-substrate intermediate. In the first mechanism, substrate radical formation leads to loss of an electron and a proton to yield a terminal olefin intermediate on one chain. Tyr459 is 4.2 Å away from the terminal carbon of the substrate and is strictly conserved, suggesting that tyrosinate might facilitate this proposed mechanism. In the second mechanism, the substrate radical might couple with the [Fe4S4]C cluster to yield an S–C bond intermediate. The sulfur atom of the [Fe4S4]C is 8.0 Å from the high-energy substrate radical intermediate. c–e, Time-dependent production of 5′-dAH (c), mAG (d) and GDGT (e) of in vitro activity assays containing either wild-type or mutant (Y459F, Y459L and M439A) forms of GDGT–MAS. These mutagenesis experiments reveal that Y459 does not have an essential role in the GDGT–MAS mechanism, which suggests that the radical intermediate is stabilized by interaction with the [Fe4S4]C cluster. The error bars represent one standard deviation for reactions conducted in triplicate, with the centre representing the mean.
