[unreadable] DESCRIPTION (provided by applicant): Arginase catalyzes the Mn(II)-dependent hydrolysis of L-arginine to produce L-ornithine and urea. In liver, this reaction is catalyzed by type I arginase and constitutes the final step of the urea cycle. Type II arginase is mitochondnal and found in tissues such as kidney, mammary gland and macrophages, where it provides a source of L-ornithine for the biosynthesis of proline and polyamines. Recent studies in this laboratory and the laboratories of our collaborators have provided compelling evidence that both type I and type Il arginases are involved in regulating the production of nitric oxide. A major goal of these studies is to dissect structure-function and structure-activity relationships for the arginase family of enzymes. We have recently determined high-resolution structures for product complexes and for complexes of the enzyme with potent boronic acid-based inhibitors. These structures are consistent with our proposed mechanism for arginase, which involves attack of a metal-bridging hydroxide on the guanidinium carbon.An expression system for the production of type I and type II arginases in E. coli has been developed for site directed mutagenesis of critical amino acids identified in the crystal structure. The origin of the exceptional substrate specificity of the enzyme will be explored through mutagenesis of residues implicated in substrate binding. The role of H141 as a proton shuttle will be evaluated through mutagenesis, chemical quench and structure determinations. Novel inhibitors of arginase will be developed and screened for potency and isozyme selectivity. Such inhibitors have therapeutic potential in the treatment of smooth muscle disorders such as erectile dysfunction. Mutagenesis studies will identify critical amino acid residues in type II arginase, and we will vigorously pursue efforts to determine the three-dimensional structure of this poorly characterized isozyme. Arginase will serve as a paradigm in structure-function analyses of the enigmatic class of Mn-metalloenzymes, including the Mn-catalases.
|Effective start/end date||07/1/02 → 06/30/08|
- National Institute of General Medical Sciences: $218,785.00
- National Institute of General Medical Sciences: $973,883.00
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