THE MOLECULAR-BASIS OF CELMERS RULES - THE STEREOCHEMISTRY OF THE CONDENSATION STEP IN CHAIN EXTENSION ON THE ERYTHROMYCIN POLYKETIDE SYNTHASE

Modular polyketide synthases (PKSs), for example, the 6-deoxyerythrono lide B synthase (DEBS) responsible for synthesis of the aglycone core of the macrolide antibiotic erythromycin, generate an impressive diver sity of asymmetric centers in their polyketide products. However, as n oted by Celmer, macrolides have the same absolute configuration at all comparable stereocenters, Understanding how the stereochemistry of ch ain extension is controlled is therefore crucial to determining the co mmon mechanism of action of these enzymes, We aimed to elucidate the m olecular basis of Celmer's rules through in vitro studies with DEBS 1- TE, a bimodular derivative of DEBS from Saccharopolyspora erythraea, w hich uses (2S)-methylmalonyl-coenzyme A to produce both D-and L-methyl centers in its triketide lactone product. We show here that condensat ion of (2S)-methylmalonyl-CoA in module 2 proceeds with decarboxylativ e inversion without cleavage of the C-H bond adjacent to the methyl gr oup; in contrast, in module 1 the chain extension process involves los s of the hydrogen attached to C-2 of the methylmalonyl-CoA precursor. The production of the D-methyl center in module 2 without loss of hydr ogen from the asymmetric center of the (2S)-methylmalonyl-CoA establis hes that condensation takes place with inversion of configuration as i n fatty acid biosynthesis, The loss of the key hydrogen from the (2S)- methylmalonyl-CoA to produce the L-methyl center generated in module 1 implies that an additional obligatory epimerization step takes place in that module. The nature and timing of the epimerization remain to b e established.