Erythromycin Biosynthesis Highly Efficient Incorporation of Polyketide Chain Elongation Intermediates into 6-Deoxyerythronolide B in an Engineered Streptomyces Host

Abstract

Feeding of (2S, 3R)-[2, 3-¹³C₂]-2-methyl-3-hydroxypentanoyl NAC thioester (1a) to the recombinant organism Streptomyces coelicolor CH999/pCK7 harboring the complete set of eryA genes from Saccharopolyspora erythraea encoding the 6-deoxyerythronolide B synthase (DEBS) resulted in the formation of 6-deoxyerythronolide B (2a) labeled with ¹³C at C-12 and C-13, as evidenced by the appearance of a pair of enhanced and coupled doublets in the ¹³C NMR spectrum. The level of ¹³C enrichment was 15-20 atom% ¹³C, as much as 100 times higher than the usually observed efficiency of incorporation of NAC thioesters into polyketide metabolites. Similar incorporation of (2S, 3R)-[3-²H, 3-¹³C]-2-methyl-3-hydroxypentanoyl NAC thioester (1b) gave 6-deoxyerythronolide B (2b) labeled with both ¹³C and deuterium at C-13. The intact incorporation of both precursors confirms the normal functioning of the recombinant DEBS proteins in the heterologous host.