Genetic Analysis of Biosynthesis of Polyketide Anthelmintic Macrolide Avermectin in Streptomyces avermitilis

Abstract

Within the 82 kb biosynthetic gene cluster for polyketide anthelmintic macrolide avermectin, the central 65 kb segment was found to be required for aglycon biosynthesis. Analysis of a 82 kb segment DNA from avermectin producer, Streptomyces avermitilis, revealed that it contains four large open reading frames (ORFs) encoding giant multifunctional polypeptides of the avermectin type-I polyketide synthase (AVES 1, AVES 2, AVES 3 and AVES 4). These clustered polyketide synthase genes responsible for avermectin biosynthesis together encode 12 homologous sets of enzyme activities (modules), each catalyzing a specific round of polyketide chain elongation and modification of β-carbon. The clustered genes encoding polyketide synthase are organized as two sets of six modular repeats, aveA1-aveA2 and aveA3-aveA4, which are convergently transcribed. The total of 55 constituent active sites were found in these four polyketide synthases but among two domains would not be functional in the process of the polyketide-chain elongation. The biosynthetic gene cluster for avermectin contains 14 additional open reading frames, some of which encode polypeptides governing other key steps in avermectin biosynthesis. Between the two sets of polyketide synthase genes lie two genes involved in postpolyketide modification. On the right of the large polyketide synthase genes is a set of genes involved in biosynthesis of methylated deoxysugar, L-oleandrose, and its transglycosylation to polyketide-derived aglycons. This cluster includes nine genes but one is not functional in the biosynthesis of avermectin. On the left side of polyketide synthase genes, Two open reading frames encoding methyltransferase and non-polyketide synthase ketoreductase involved in post- polyketide modification are located and an adjacent gene encodes a regulatory function which may be involved in activation of the transcription of avermectin biosynthetic genes.