Cerulenin resistance in a cerulenin-producing fungus. III. Studies on active-site peptides of fatty acid synthetase from Cephalosporium caerulens.
Active-site peptides of acetyl transferase, condensing enzyme and acyl carrier protein in the neighborhood of the prosthetic group, 4'-phosphopantetheine, of Cephalosporium caerulens fatty acid synthetase were investigated. The enzyme was reacted with [14C]acetyl-CoA or [14C]iodoacetamide. 14C-Labeled enzyme was digested with pepsin, trypsin or both. 14C-Labeled peptides were isolated by several purification procedures. The amino acid sequence of the active site of condensing enzyme was determined to be Tyr-Gln-Val-Glu-Ser-Cys-Pro-Ile-Leu-Glu-Gly-Lys and that of acetyl transferase was Phe-Ser-Gly-Ala-Thr-Gly-His-Ser-Gln-Gly. The amino acid composition around the 4'-phosphopantetheine-carrying serine was determined to be Asx2, Thr, Ser, Glx3, Gly2, Ala, Ile, Leu3, and Lys. When these active-site peptides were compared with those of Saccharomyces cerevisiae synthetase, a high degree of homology was observed in the active-site peptides of the acetyl transferase and acyl carrier protein domains. However, that of the condensing enzyme domain gave lower homology. These findings may support the assumption that the low reactivity of cerulenin with C. caerulens synthetase is a consequence of the structure of the condensing enzyme domain.
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000164:Acremonium / Q000201:enzymology*; D000595:Amino Acid Sequence; D001665:Binding Sites; D002250:Carbon Radioisotopes; D002569:Cerulenin / Q000096:biosynthesis / Q000633:toxicity; D004351:Drug Resistance; D064429:Fatty Acid Synthases / Q000378:metabolism*; D007460:Iodoacetamide / Q000378:metabolism; D007700:Kinetics; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding
D002250:Carbon Radioisotopes; D010446:Peptide Fragments; D002569:Cerulenin; D000105:Acetyl Coenzyme A; D064429:Fatty Acid Synthases; D007460:Iodoacetamide