New insights into the first oxidative phenol coupling reaction during vancomycin biosynthesis

Label from 18O 2 is not incorporated into the monocyclic product shown during catalysis by OxyB. Also, a model hexapeptide containing ( R)-Tyr6, instead of ( S)-Tyr6, is converted by OxyB into the corresponding epimeric monocyclic product. OxyB catalyzes the first oxidative phenol coupling reaction in vancomycin biosynthesis. OxyB is a P450 hemoprotein whose activity is strictly dependent upon the presence of molecular oxygen. Here, it was shown that label from 18O 2 is not incorporated into the monocyclic product during catalysis by OxyB. In addition, it was shown that OxyB can convert a model hexapeptide substrate containing ( R)-Tyr6, instead of ( S)-Tyr6, covalently linked as a C-terminal thioester to a peptidyl carrier protein (PCP-7S) derived from the vancomycin non-ribosomal peptide synthetase (NRPS), into the corresponding epimeric monocyclic product. The binding of this epimeric hexapeptide-PCP conjugate to the Fe(III) form of OxyB, as monitored by UV–vis spectroscopy, revealed a K d = 35 ± 5 μM. Thus, the enzyme reveals a surprising lack of stereospecificity in the binding and transformation of these epimeric substrates.