Biosynthesis of the 4-Methyloxazoline-Containing Nonribosomal Peptides, JBIR-34 and -35, in Streptomyces sp. Sp080513GE-23

Abstract

JBIR-34 and -35 produced by Streptomyces sp. Sp080513GE-23 are nonribosomal peptides that possess an unusual 4-methyloxazoline moiety. Through draft genome sequencing, cosmid cloning, and gene disruption, the JBIR-34 and -35 biosynthesis gene cluster (fmo cluster) was identified; it encodes 20 proteins including five nonribosomal peptide synthetases (NRPSs). Disruption of one of these NRPS genes (fmoA3) resulted in no JBIR-34 and -35 production and accumulation of 6-chloro-4-hydroxyindole-3-carboxylic acid. Stable isotope-feeding experiments indicated that the methyl group of the methyloxazoline ring is derived from alanine rather than methionine. A recombinant FmoH protein, a glycine/serine hydroxymethyltransferase homolog, catalyzed conversion of α-methyl-l-serine into d-alanine (the reverse reaction of α-methyl-l-serine synthesis catalyzed by FmoH in vivo). Taken together, we concluded that α-methyl-l-serine synthesized from d-alanine is incorporated into JBIR-34 and -35 to form the 4-methyloxazoline moiety. We also propose the biosynthesis pathway of JBIR-34 and -35.

Metabologenomics: Correlation of Microbial Gene Clusters with Metabolites Drives Discovery of a Nonribosomal Peptide with an Unusual Amino Acid Monomer

“…Tambromycin’s distinguishing features are a hydroxylated and chlorinated indole, a methyl-oxazoline, and 2-methyl-serine, substructures also found in JBIR-34/35 ( Figure 2 A). 10 However, an unusual pyrrolidine-containing amino acid was found upon complete structural elucidation of tambromycin by NMR, also recently reported by Izumikawa et al under the name JBIR-126. 11 Here we report the absolute stereochemistry of the tambroline monomer, the structures of tambromycin and two of its analogues, and the biosynthetic gene cluster for the compound, shown to be expressed in nine different strains.…”

Pyrrolidine-containing peptides, JBIR-126, -148, and -149, from Streptomyces sp. NBRC 111228

“…The established methyl oxazoline residue was also supported by comparison of the chemical shifts with those of JBIR-34. 8,9 The connectivity among these four units was established by the 1 H-13 C long-range couplings from H-2 to C-1 0 , from the a-methine proton H-2 00 and the amide proton 2 00 -NH to the amide carbonyl carbon C-5 0 , from the a-methine proton H-2 00 and the amide proton 2 000 -NH to an amide carbonyl carbon C-1 00 (d C 167.6). Finally, according to the molecular formula and the 13 C chemical shift of C-6, a chlorine atom was introduced at C-6.…”

“…8 The unique structures of JBIR-34 and -35 have been biosynthesized by a novel NRPS gene resided in Sp080513GE-23. 9 Biosynthetic studies of compounds 1-3 will be reported elsewhere. …”

Unrivalled diversity: the many roles and reactions of bacterial cytochromes P450 in secondary metabolism

“…229 An example of a two-step transformation of a PCP-bound amino acid has been suggested to occur in the biosynthesis of the cyclic peptides JBIR-34 and JBIR-35. 230 In this pathway, the di-domain NRPS module FmoA1 presents a tryptophan residue (or chlorinated derivative) to the P450 FmoC, which performs two oxidation reactions; hydroxylation of the indole ring as well as b-hydroxylation.…”