Identification of DEBS 1, DEBS 2 and DEBS 3, the multienzyme polypeptides of the erythromycin‐producing polyketide synthase from Saccharopolyspora erythraea

The ery A region of the erythromycin biosynthetic gene cluster of Saccharopolyspora erythraea has previously been shown to contain three large open reading frames (ORFs) that encode the components of 6‐deoxyerythronolide B synthase (DEBS). Polyclonal antibodies were raised against recombinant proteins obtained by overexpression of 3′ regions of the ORF2 and ORF3 genes. In Western blotting experiments, each antiserum reacted strongly with a different high molecular weight protein in extracts of erythromycin‐producing S. erythraea cells. These putative DEBS 2 and DEBS 3 proteins were purified and subjected to N‐terminal sequence analysis. The protein sequences were entirely consistent with the translation start sites predicted from the DNA sequences of ORFs 2 and 3. A third high molecular weight protein co‐purified with DEBS 2 and DEBS 3 and had an N‐terminal sequence that matched a protein sequence translated from the DNA sequence some 155 base pairs upstream from the previously proposed start codon of ORF1.