A three enzyme pathway for 2-amino-3-hydroxycyclopent-2-enone formation and incorporation in natural product biosynthesis

Article abstract of DOI:10.1021/ja1002845

A number of natural products contain a 2-amino-3-hydroxycyclopent-2-enone five membered ring, termed C₅N, which is condensed via an amide linkage to a variety of polyketide-derived polyenoic acid scaffolds. Bacterial genome mining indicates three tandem ORFs that may be involved in C₅N formation and subsequent installation in amide linkages. We show that the protein products of three tandem ORFs (ORF33-35) from the ECO-02301 biosynthetic gene cluster in Streptomyces aizunenesis NRRL-B-11277, when purified from Escherichia coli, demonstrate the requisite enzyme activities for C₅N formation and amide ligation. First, succinyl-CoA and glycine are condensed to generate 5-aminolevulinate (ALA) by a dedicated PLP-dependent ALA synthase (ORF34). Then ALA is converted to ALA-CoA through an ALA-AMP intermediate by an acyl-CoA ligase (ORF35). ALA-CoA is unstable and has a half-life of ～10 min under incubation conditions for off-pathway cyclization to 2,5-piperidinedione. The ALA synthase can compete with the nonenzymatic decomposition route and act in a novel second transformation, cyclizing ALA-CoA to C₅N. C ₅N is then a substrate for the third enzyme, an ATP-dependent amide synthetase (ORF33). Using octatrienoic acid as a mimic of the C₅₆ polyenoic acid scaffold of ECO-02301, formation of the octatrienyl-C ₅N product was observed. This three enzyme pathway is likely the general route to the C₅N ring system in other natural products, including the antibiotic moenomycin.