Valine dehydrogenase (VDH) from Streptomyces hygroscopicus CH-7 was purified 327-fold with 21% yield, using ion exchange and hydrophobic chromatography. The enzyme had an Mr 31000 in denaturing conditions and an Mr 63000 in gel filtration chromatography, indicating that it is homo-dimer. The most preferable substrates are L-valine in deamination reaction and 2-ketoisovalerate in amination reaction. The enzyme requires NAD⁺ as a cofactor. The VDH from S. hygroscopicus CH-7 shows maximum activity at approximately pH 10.7 and 9.7 for deamination and amination reactions, respectively. The enzyme was significantly inhibited by p-chloromercuri-benzoate, Hg²⁺ and other metal ions, which suggests that the presence of SH- groups are necessary for the catalytic reaction. The apparent Michaelis constants for L-valine, NAD⁺ 2-ketoisovalerate, NADH and NH4⁺ were: 1.26 mM, 0.164 mM, 0.41 mM, 0.026 mM and 50.1 mM.

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The effect of the upstream region of the β-lactamase structural gene was studied on the production of β-lactamase from Streptomyces fradiae Y59. In contrast to the case in S. cacaoi, β-lactamase activity was weakly enhanced by the presence of the upstream 5-kb DNA fragment but was not induced by 7-ACA. The sequence analysis of this region revealed a couple of putative regulatory proteins, although their actual functions were not clear. It was therefore suggested that at least one of the ORFs might be involved in the weak enhancement of β-lactamase gene expression of S. fradiae Y59

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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.

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The gene cluster encoding biosynthesis of the polyketide backbone of erythromycin is organised as repeated modules of functional domains. The similarity of the domain arrangements among the complex polyketide natural products provides a strategy to generate an array of different structures using a combinatorial approach. More than fifty 6-deoxyerythronolide B (6dEB) analogs were obtained by substitution of catalytic domains via single, double and triple alterations. This genetic manipulation approach to engineer such molecules, which would otherwise be impractical to produce by chemical methods, is very challenging and promising for the development of novel polyketides that could have utility in pharmaceutical applications in the future.

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