Molecular biology of polyketide biosynthesis

Streptomyces species and related genera synthesize a large number of second ary metabolites, many of which are biologically active. Amongst them, polyk etides is the largest class. Polyketides are a structurally diverse family of natural products with a broad range of biological activities. The format ion of polyketides is very similar to the biosynthesis of long chain fatty acids - both in the enzymatic reactions that take place and the enzyme prot eins that are involved. During the last decade many polyketide gene-cluster s have been cloned and sequenced. DNA sequencing has shown that the cluster s have substantial homology suggesting that they originated from a common a ncestor. This similarity has resulted in the development of combinatorial b iology techniques to create novel chemical entities. Two approaches have be en used: targeted manipulation, e.g. disruption and, often, replacement of certain genes involved in the biosynthetic pathway, and the random approach , e.g. "gene shuffling". A targeted approach has been used to generate seve ral novel scaffolds by manipulation of the S. rimosus oxytetracycline gene- cluster. Genes encoding ketosynthase, alpha and beta, ketoreductase, cyclas e/aromatase and C-6 hydroxylase were disrupted to construct four recombinan t strains. Thin layer chromatography and high pressure liquid chromatograph y of extracts from fermentation broths of all four recombinants showed that they produce about 20 potentially novel metabolites, 12 of which have been characterised chemically. In this review, data on the disruption and repla cement of the otcC gene will be described in more detail.