The conversion of a dormant bacterial (Bacillus) spore into a vegetative bacterium by the process of spore germination is a relatively simple differentiating system which is readily amenable to biochemical analysis. A major feature of this differentiation process is extensive and rapid hydrolysis of spore protein. Degradative enzymes are important in regulating enzyme levels, especially in mammalian cells, and in several cases specific proteolytic enzymes have been shown to be involved in regulation of several processes including bacterial sporulation, yeast budding, and sea urchin egg development. Therefore, investigation of the role of degradative enzymes in spore germination may provide insights into the control of differentiation in higher organisms. The objective of this project is the elucidation of the mechanism of action and the function of proteolytic enzymes in the germination of bacterial spores - especially with regard to potential control function.
Specific aims are to: 1) obtain reproducible translation of the B protein in vitro; 2) clone the genes for the A, C and possibly B proteins; 3) use the cloned genes for the A, B and C proteins to determine the organization and sequence of these genes; 4) use the cloned genes as probes to identify the site of synthesis (mother cell or forespore) of the rRNAs for the A, B and C proteins; 5) use the cloned B. megaterium genes as probes in attempts to isolate the comparable genes from B. subtilis and B. cereus; and 6) use the newly developed radioimmunoassay for the spore protease to study its synthesis, localization, degradation and regulation.
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