Covering: 2002–2005 This review covers advances in understanding of the biosynthesis of polyketidesproduced by type II PKS systems at the genetic, biochemical and structural levels.

The saccharomicins A and B, produced by the actinomycete Saccharothrix espanaensis , are oligosaccharide antibiotics. They consist of 17 monosaccharide units and the unique aglycon N -( m , p -dihydroxycinnamoyl)taurine. To investigate candidate genes responsible for the formation of trans-m , p -dihydroxycinnamic acid (caffeic acid) as part of the saccharomicin aglycon, gene expression experiments were carried out in Streptomyces fradiae XKS. It is shown that the biosynthetic pathway for trans -caffeic acid proceeds from l -tyrosine via trans-p -coumaric acid directly to trans -caffeic acid, since heterologous expression of s am8 , encoding a tyrosine ammonia-lyase, led to the production of trans-p -hydroxycinnamic acid (coumaric acid), and coexpression of sam8 and sam5 , the latter encoding a 4-coumarate 3-hydroxylase, led to the production of trans-m , p -dihydroxycinnamic acid. This is not in accordance with the general phenylpropanoid pathway in plants, where trans-p -coumaric acid is first activated before the 3-hydroxylation of its ring takes place.

The entire simocyclinone biosynthetic cluster ( sim gene cluster) from the producer Streptomyces antibioticus Tü6040 was identified on six overlapping cosmids (1N1, 5J10, 2L16, 2P6, 4G22, and 1K3). In total, 80.7 kb of DNA from these cosmids was sequenced, and the analysis revealed 49 complete open reading frames (ORFs). These ORFs include genes responsible for the formation and attachment of four different moieties originating from at least three different pools of primary metabolites. Also in the sim gene cluster, four ORFs were detected that resemble putative regulatory and export functions. Based on the putative function of the gene products, a model for simocyclinone D8 biosynthesis was proposed. Biosynthetic mutants were generated by insertional gene inactivation experiments, and culture extracts of these mutants were analyzed by high-performance liquid chromatography. Production of simocyclinone D8 was clearly detectable in the wild-type strain but was not detectable in the mutant strains. This indicated that indeed the sim gene cluster had been cloned.

Analysis of the simocyclinone biosynthesis (sim) gene cluster of Streptomyces antibioticus Tü6040 led to the identification of a putative pathway specific regulatory gene simReg1. In silico analysis places the SimReg1 protein in the OmpR-PhoB subfamily of response regulators. Gene replacement of simReg1 from the S. antibioticus chromosome completely abolishes simocyclinone production indicating that SimReg1 is a key regulator of simocyclinone biosynthesis. Results of the DNA-shift assays and reporter gene expression analysis are consistent with the idea that SimReg1 activates transcription of simocyclinone biosynthesis, transporter genes, regulatory gene simReg3 and his own transcription. The presence of extracts (simocyclinone) from S. antibioticus Tü6040 × pSSimR1-1 could dissociate SimReg1 from promoter regions. A preliminary model for regulation of simocyclinone biosynthesis and export is discussed.

The gram-positive bacterium Streptomyces aureofaciens Tü117 produces the acyclic polyene antibiotic α-lipomycin. The entire biosynthetic gene cluster ( lip gene cluster) was cloned and characterized. DNA sequence analysis of a 74-kb region revealed the presence of 28 complete open reading frames (ORFs), 22 of them belonging to the biosynthetic gene cluster. Central to the cluster is a polyketide synthase locus that encodes an eight-module system comprised of four multifunctional proteins. In addition, one ORF shows homology to those for nonribosomal peptide synthetases, indicating that α-lipomycin belongs to the classification of hybrid peptide-polyketide natural products. Furthermore, the lip cluster includes genes responsible for the formation and attachment of d -digitoxose as well as ORFs that resemble those for putative regulatory and export functions. We generated biosynthetic mutants by insertional gene inactivation. By analysis of culture extracts of these mutants, we could prove that, indeed, the genes involved in the biosynthesis of lipomycin had been cloned, and additionally we gained insight into an unusual biosynthesis pathway.