A free-standing condensation enzyme catalyzing ester bond formation in c-1027 biosynthesis

Academic Article
uri icon

publication date

  • 2009

abstract

  • Nonribosomal peptide synthetases (NRPSs) catalyze the biosynthesis of many biologically active peptides and typically are modular, with each extension module minimally consisting of a condensation, an adenylation, and a peptidyl carrier protein domain responsible for incorporation of an amino acid into the growing peptide chain. C-1027 is a chromoprotein antitumor antibiotic whose enediyne chromophore consists of an enediyne core, a deoxy aminosugar, a benzoxazolinate, and a beta-amino acid moiety. Bioinformatics analysis suggested that the activation and incorporation of the beta-amino acid moiety into C-1027 follows an NRPS mechanism whereby biosynthetic intermediates are tethered to the peptidyl carrier protein SgcC2. Here, we report the biochemical characterization of SgcC5, an NRPS condensation enzyme that catalyzes ester bond formation between the SgcC2-tethered (S)-3-chloro-5-hydroxy-beta-tyrosine and (R)-1-phenyl-1,2-ethanediol, a mimic of the enediyne core. SgcC5 uses (S)-3-chloro-5-hydroxy-beta-tyrosyl-SgcC2 as the donor substrate and exhibits regiospecificity for the C-2 hydroxyl group of the enediyne core mimic as the acceptor substrate. Remarkably, SgcC5 is also capable of catalyzing amide bond formation, albeit with significantly reduced efficiency, between (S)-3-chloro-5-hydroxy-beta-tyrosyl-(S)-SgcC2 and (R)-2-amino-1-phenyl-1-ethanol, an alternative enediyne core mimic bearing an amine at its C-2 position. Thus, SgcC5 is capable of catalyzing both ester and amide bond formation, providing an evolutionary link between amide- and ester-forming condensation enzymes.