Exploiting a C–N Bond Forming Cytochrome P450 Monooxygenase for C–S Bond Formation

C–S bond formation reactions are widely distributed in the biosynthesis of biologically active molecules, and thus have received much attention over the past decades. Herein, we report intramolecular C–S bond formation by a P450 monooxygenase, TleB, which normally catalyzes a C−N bond formation in teleocidin biosynthesis. Based on the proposed reaction mechanism of TleB, a thiol‐substituted substrate analogue was synthesized and tested in the enzyme reaction, which afforded the unprecedented sulfur‐containing thio‐indolactam V, in addition to an unusual indole‐fused 6/5/8‐tricyclic product whose structure was determined by the crystalline sponge method. Interestingly, conformational analysis revealed that the SOFA conformation is stable in thio‐indolactam V, in sharp contrast to the major TWIST form in indolactam V, resulting in differences in their biological activities. Cytochrome P450 monooxygenases catalyze a wide range of oxidative transformations. TleB, a P450 monooxygenase involved in the biosynthesis of teleocidin, catalyzes C–S bond formation through both oxidation‐mediated and radical‐mediated mechanisms to generate a sulfur‐containing indolactam and an unusual 6/5/8 tricyclic compound. These results may enable the production of new unnatural compounds using enzymes as biocatalysts.