A polyketide synthase in glycopeptide biosynthesis - The biosynthesis of the non-proteinogenic amino acid (S)-3,5-dihydroxyphenylglycine

Balhimycin, a vancomycin-type antibiotic from Amycolatopsis mediterranei, c ontains the unusual amino acid (S)-3,5-dihydroxyphenylglycine (Dpg), with a n acetate-derived carbon backbone. After sequence analysis of the biosynthe tic gene cluster, one gene, dpgA, for a predicted polyketide synthase (PKS) was identified, sharing 20-30% identity with plant chalcone synthases. Ina ctivation of dpgA resulted in loss of balhimycin production, and restoratio n was achieved by supplementation with 3,5-dihydroxyphenylacetic acid, whic h is both a possible product of a PKS reaction and a likely precursor of Dp g. Enzyme assays with the protein expressed in Streptomyces lividans showed that this PKS uses only malonyl-CoA as substrate to synthesize 3,5-dihydro xyphenylacetic acid. The PKS gene is organized in an operon-like structure with three downstream genes that are similar to enoyl-CoA-hydratase genes a nd a dehydrogenase gene. The heterologous co-expression of all four genes l ed to accumulation of 3,5-dihydroxy-phenylglyoxylic acid. Therefore, we now propose a reaction sequence. The final step in the pathway to Dpg is a tra nsamination. A predicted transaminase gene was inactivated, resulting in ab olished antibiotic production and accumulation of 3,5-dihydroxyphenylglyoxy lic acid. Interestingly, restoration was only possible by simultaneous supp lementation with (S)-3,5-dihydroxyphenylglycine and (S)-4-hydroxyphenylglyc ine, indicating that the transaminase is essential for the formation of bot h amino acids.