Many naturally occuring strains of bacteria produce antibiotic substances. Production of such substances may play an important role in a strain's ability to become established in a particular niche. A class of low-molecular-weight antibiotic substances produced by enteric bacteria was discovered in Spain in 1975 and termed """"""""microcins"""""""". The studies that have followed have shown that among clinical isolates microcins are very common. One particular microcin, B17, inhibits bacterial DNA replication. It has been the subject of careful genetic analyses and the results thus far indicate that its production is the result of a rather complex pathway. The genetic and biochemical experiments proposed will analyze in detail the regulation of synthesis and mode of action of the peptide antibiotic microcin B17 produced by E. coli. The microcin will be purified and characterized. This will be followed by the isolation of structural gene mutations to obtain altered forms of the molecule. In vitro experiments will then help in elucidating the molecular interactions between the peptide and the replication machinery of the cell which lead to inhibition of synthesis. The pathway leading to the production of active microcin will also be studied through the isolation of mutations in all the genes involved in the process. Microcin B-17, like most antibiotics, is only produced in stationary phase. Through the use of gene fusions and the isolation of mutants defective in regulation of expression the molecular mechanism of this regulation will be analyzed. Transcription antitermination has been implicated in stationary phase regulation and experiments are designed to critically test this hypothesis.
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