Streptomyces species synthesize more than 70 medically useful antibioties as well as antitumor and antiparasitic agents. These compounds are synthesized from primary metabolites during nutritional limitation of growth and the initiation of sporulation. Surprisingly, very little is known about primary metabolism or the sechanisms that trigger antibiotic production and sporulation in these bacteria. In order to increase our understanding of Streptomyces basic physiology, the regulation of glutamine synthetase (GS), a key enzyme in ammoniin assimilation, is being studied in S. coelicolor. Analysis of the cloned I. coelicolor GS structural gene (sm) shows that transcription of the SM promoter is nitrogen regulated during vegetative growth. Two approaches are being used to identify tbe factors that mediate this regulation. First, nucleotide regions required for the regulation and expression of the LM pronoter are being identified by constructing glnA-xylE transcriptional fusions, and isolating nutations that alter expression and regulation of the glnA-xylE fusions. Secondly, glnA regulatory factors are being genetically identified by isolating mutants with altered expression of the glnA promoters. The glutamine-requiring ClassI GlnR mutants are unable to transcribe the nitrogen-regulated glnA promoter. The defective gene product in this nutant has been cloned by complementation and is being identified subcloned to facilitate DNA sequencing. Since expression of the nitrogen-regulated enzyme urease is also deficient in these mutants, a global nitrogen regulatory system may be present in .1. coelicolor. The cloned &W DNA will be used to study interactions between the &W promoters and the genetically identified glnA regulatory proteins, RNA polymerase and small nitrogencontaining metabolites. An understanding of nitrogen metabolism in 5. coelicolor should facilitate increased antibiotic production by genetic manipulation of primary uetabolite pools, or by choice of the appropriate growth nedium. Furthermore, since primary metabolism impinges on both antibiotic production and sporulation in Streptomyces, these studies may help define mechanisms that regulate these processes in Streptomyces.
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