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 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.
Caffrey
P
P
Department of Biochemistry, University of Cambridge, UK.
Bevitt
D J
DJ
Staunton
J
J
Leadlay
P F
PF
D000595:Amino Acid Sequence; D004917:Erythromycin / Q000378:metabolism*; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000302:isolation & purification; D010455:Peptides / Q000737:chemistry*; D016930:Saccharopolyspora / Q000201:enzymology*; D012689:Sequence Homology, Nucleic Acid
D009097:Multienzyme Complexes; D010455:Peptides; D004917:Erythromycin