The long range objective of the proposed research is to discover new mechanisms of regulation of enzyme activity, mechanisms that operate in addition to modification of enzyme activity by non-covalent binding of effector molecules and repression of enzyme synthesis. In particular, we are seeking control mechanisms that may be related to enzyme inactivation and turnover processes that may be important during development and differentiation. The specific experimental system to be analyzed is the biochemical fate of the enzymes of de novo pyrimidine and purine biosynthesis during growth and sporulation of Bacillus subtilis. These synthetic capacities are lost during endospore formation. In the case of de novo pyrimidine synthesis, aspartic transcarbamylase has been shown to be rapidly inactivated at the end of log phase growth, in a process that requires metabolic energy, but which is not blocked by antibiotics that inhibit protein synthesis. Immunochemical studies indicate that cross-reactive protein is lost in parallel with activity, which suggests turnover follows inactivation. In this case of purine biosynthesis, glutamine phosphoribosylpyrophosphate amidotransferase has been shown to be activated also, but by a process that requires molecular oxygen, rather than metabolic energy. This process can be reconstructed in vitro and is regulated by ligands of the enzyme. This proposal contemplates learning the biochemical nature of these inactivation processes and/or turnover processes by a variety of physiological, biochemical and immunochemical techniques. BIBLIOGRAPHIC REFERENCES: Robert L. Switzer, "The Inactivation of Microbial Enzymes in Vivo," Ann. Rev. Microbiol., in press (1977). M. R. Maurizi and R. L. Switzer, "Immunochemical Studies of the Inactivation of Aspartate Transcarbamylase in Bacillus subtilis, Federation Proc., in press (1977) (abstract).