This proposal seeks support for an investigation of the biochemistry and molecular genetics of macrolide and polyether antibiotic production by streptomycetes as a means to facilitate the study of the enzymology of these antibiotics' biosynthesis in the future. Two clinically and commercially important antibiotics, erythromycin A and lasalocid A, are the focal points of this research. The principal long range goal is to clone the gene(s) coding the enzymatic capability of macrolactone or alicyclic carbon chain formation. Methods already developed by other scientists for the artificial and natural genetic manipulation of actinomycetes will be applied to gene cloning and amplification in S. erythreus and S. lasaliensis to increase the cellular titer of specified antibiotic synthetases. Subsidiary long range goals will be a resolution of the question of the location of the antibiotic genes (chromosome vs. plasmid), an investigation of the natural regulatory systems for antibiotic gene expression, and the analysis of the potential for hybrid or unnatural antibiotic formation - all studied by further development of recombinant DNA methods in streptomycetes. Research endeavours of lesser importance that will arise during execution of the latter are the question of heterologous DNA replication and expression (S. erythreus in S. lividans or E. coli) and the value of site-directed mutagenesis as a tool for probing antibiotic biosynthesis.
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