(ET), seems to be crucial for efficient desaturation. Thus, perturbing the major PT chains is expected to
impair the native reaction and (potentially) amplify minor reaction channels, such as the substrate
hydroxylation. To verify this hypothesis, we mutated the four residues mentioned above into their counterparts present in a soluble methane monooxygenase (sMMO), and determined the reaction products of
mutants. We found that the mutations significantly promote residual monohydroxylation activities on
stearoyl-CoA, often at the expense of native desaturation activity. The favored hydroxylation positions
are C9, followed by C10 and C11. Reactions with unsaturated substrate, oleoyl-CoA, yield erythro-9,10-
... More
(ET), seems to be crucial for efficient desaturation. Thus, perturbing the major PT chains is expected to
impair the native reaction and (potentially) amplify minor reaction channels, such as the substrate
hydroxylation. To verify this hypothesis, we mutated the four residues mentioned above into their counterparts present in a soluble methane monooxygenase (sMMO), and determined the reaction products of
mutants. We found that the mutations significantly promote residual monohydroxylation activities on
stearoyl-CoA, often at the expense of native desaturation activity. The favored hydroxylation positions
are C9, followed by C10 and C11. Reactions with unsaturated substrate, oleoyl-CoA, yield erythro-9,10-
diol, cis-9,10-epoxide and a mixture of allylic alcohols. Additionally, using 9- and 11-hydroxystearoylCoA, we showed that the desaturation reaction can proceed only with the hydroxyl group at position
C11, whereas the hydroxylation reaction is possible in both cases, i.e. with hydroxyl at position C9 or
C11. Despite the fact that the overall outcome of hydroxylation is rather modest and that it is mostly
the desaturation/hydroxylation ratio that is affected, our results broaden understanding of the origin
of chemo- and stereoselectivity of the D9
D and provide further insight into the catalytic action of the
NHFe2 enzymes
