ERYTHROMYCIN BIOSYNTHESIS - HIGHLY EFFICIENT INCORPORATION OF POLYKETIDE CHAIN ELONGATION INTERMEDIATES INTO 6-DEOXYERYTHRONOLIDE-B IN AN ENGINEERED STREPTOMYCES HOST

Feeding of (2S,3R)-[2,3-C-13(2)]-2-methyl-3-hydroxypentanoyl NAC thioe ster (1a) to the recombinant organism Streptomyces coelicolor CH999/pC K7 harboring the complete set of eryA genes from Saccharopolyspora ery thraea encoding the 6-deoxyerythronolide B synthase (DEBS) resulted in the formation of 6-deoxyerythronolide B (2a) labeled with C-13 at C-1 2 and C-13, as evidenced by the appearance of a pair of enhanced and c oupled doublets in the C-13 NMR spectrum. The level of C-13 enrichment was 15 similar to 20 atom% C-13, as much as 100 times higher than the usually observed efficiency of incorporation of NAC thioesters into p olyketide metabolites. Similar incorporation of (2S,3R)-[3-H-2,3-C-13] -2-methyl-3-hydroxypentanoyl NAC thioester (1b) gave 6-deoxyerythronol ide B (2b) labeled with both C-13 and deuterium at C-13. The intact in corporation of both precursors confirms the normal functioning of the recombinant DEBS proteins in the heterologous host.