Genetic studies on a soil streptomyces sp. that produces an antifungal compoud

Abstract

In an effort to identify novel antifungal compounds, soil isolates from different parts of Singapore were screened and several isolates had been identified in an earlier study. One such isolate was 98- 62. The aim of the current study is to study the genes responsible for the biosynthesis of the antifungal compound produced by the soil isolate 98-62. As several of the antifungal compounds synthesized by bacterial strains are encoded by PKS I genes, it was postulated that the antifungal compound produced by the soil isolate 98-62 could also be encoded by PKS I genes. When the genomic DNA of the soil isolate 98- 62 was probed with PKS I specific genes, strong multiple hybridizing bands were observed as is characteristic of PKS I genes. Furthermore, thin layer chromatography separation of the antifungal compound produced by the soil isolate 98-62 compared to the Rf values of complex polyketides rapamycin and FK506. Complex polyketides are encoded by PKS I genes. By homology based approach, a total of 11.6 kb of contiguous PKS I genes were then cloned from the soil isolate 98- 62 and characterized. Sequence analysis of the cloned genes confirmed that a partial cluster of PKS I genes, spanning 2 partial open reading frames, encompassing one complete and two partial modules of PKS I genes had been cloned. To prove the functional involvement of the cloned PKS I genes in the biosynthesis of the antifungal compound, gene disruption using four different gene fragments of the cloned PKS I genes were carried out. Gene disruption experiment using internal fragments of the cloned PKS I genes abolished the antifungal compound production. This work provides functional evidence that the antifungal compound produced by the soil isolate 98- 62 is encoded by the cloned partial PKS I cluster of genes.