Dinuclear Iron Cluster‐Containing OxygenaseCmlA

Abstract

AbstractCmlA is a tailoring enzyme from the nonribosomal peptide synthetase (NRPS)‐based biosynthetic pathway for chloramphenicol expressed inStreptomyces venezuelae. The enzyme is a monooxygenase that catalyzes the β‐hydroxylation ofl‐para‐aminophenylalanine (l‐PAPA) covalently tethered to the NRPS CmlP. Spectroscopic and X‐ray crystallographic studies reveal an oxo‐bridged dinuclear iron cluster in the active site that binds and activates O2. The structure shows that the subunits of the homodimeric CmlA are composed of a 248 residue N‐terminal domain with a novel fold and a 284 residue C‐terminal domain with a metallo‐β‐lactamase (MBL) fold where the diiron cluster is bound. Oxygenase activity has not been previously observed for a diiron cluster in an MBL fold and no other diiron cluster enzyme is known to catalyze β‐hydroxylation. In the diferrous state, the diiron cluster is reactive with O2, but the reaction is very slow in the absence of the CmlP–l‐PAPA complex. Thus, CmlA is regulated so that O2is activated only when substrate is bound. CmlA is the first diiron cluster‐containing enzyme known to be active in an NRPS pathway, but a gene search reveals numerous homologs of CmlA that are likely to catalyze similar reactions during biosynthesis of other natural products.