Biosynthesis of acylpeptidolactones of the daptomycin type. A comparative analysis of peptide synthetases forming A21978C and A54145.
A21978C and A54145 are antibacterial 13-residue peptides with a medium-chain-acylated amino terminus and a 10-residue lactone ring; they are produced by strains of Streptomyces roseosporus and Streptomyces fradiae, respectively. The structural differences in their peptide chains, which include amino acid replacements and modifications (L-Glu2-->L-Asn, L-Asn(OH)3-->L-Asp, Sar5-->Gly, L-Ala6-->L-Orn, L-Lys8-->D-Ala, L-Asp(OMe)9-->L-Asp, L-Asn11-->D-Ser, and L-lle13-->L-Kyn; Sar = sarcosine; L-Orn = L-ornithine, L-Kyn = L-kynurenine), reside in the multienzymatic templates directing their biosynthesis. We have examined the peptide synthetases employing immunodetection and substrate activation detected by the amino-acid-dependent ATP-PP1-exchange reaction. Two different antibodies specific for actinomycin synthetase 2 and a peptide sequence characteristic of acyl-CoA-synthetases/peptide synthetases were applied. For the A21978 system two peptide synthetases of 670 and 240 kDa were detected, together with two similar proteins of 630 and 440 kDa occurring in varying amounts. The latter are suggested to be degradation products of an unstable multienzyme. Activation of L-Asp, L-Thr, Gly, L-Orn, L-Ala and L-Ser were assigned to the high-molecular-mass components of 670, 630 and 440 kDa. The 240-kDa protein was purified to homogeneity and shown to catalyse activation of L-kynurenine. The A54145 system consisted of three peptide synthetases of 690, 590 and 250 kDa. Activations of L-Asn. L-Thr and Gly were found. The 250-kDa synthetase was capable of activating isoleucine and valine. Both systems thus show a comparable organisation; implications for the modular construction of their peptide synthetases are discussed.
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Institut für Biochemie und Molekulare Biologie, Technische Universität Berlin, Germany.
von Döhren
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D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D003429:Cross Reactions; D036341:Intercellular Signaling Peptides and Proteins; D008074:Lipoproteins / Q000096:biosynthesis; D008970:Molecular Weight; D010453:Peptide Synthases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D010455:Peptides / Q000276:immunology; D013302:Streptomyces / Q000201:enzymology*
C065208:A54145; D000900:Anti-Bacterial Agents; D036341:Intercellular Signaling Peptides and Proteins; D008074:Lipoproteins; D010455:Peptides; C044687:A 21978C1; D010453:Peptide Synthases