This is a request for an ADAMHA Research Scientist Award. Proteinases play an essential role in normal brain function by catalyzing the synthesis and degradation of neuropeptides. Abnormal proteolysis has been proposed to underlie pathological changes in brain such as the deposition of beta- amyloid in Alzheimer's disease and in Down's syndrome. We wish to continue and extend our studies on brain and pituitary proteinases. A unique multicatalytic proteinase complex (MPC) discovered and characterized in our laboratory a decade earlier, has now become the focus of intensive worldwide investigation. MPC recognized as the major extra-lysosomal protein degrading system and the only known proteinase with multiple active sites, is present in all eukaryotic cells. We recently discovered a co- purifying cAMP-dependent protein kinase that appears functionally linked to MPC. We propose exploration of the possible role of MPC in the generation of beta-amyloid, detailed characterization of the substrate specificities of each of the components, synthesis of MPC inhibitors, and studies on the nature of the subunits of MPC. We demonstrated that the two pyroglutamyl peptidases involved in the degradation of thyrotropin releasing hormone (TRH) are subject to multiple modes of regulation. The molecular cloning of pyroglutamyl peptidase II, the first characterized neuropeptide specific peptidase will be conducted in collaborative studies, enabling us to study regulation of this enzyme at the transcriptional level. The biological activity of the brain renin-angiotensin system may be expressed by either angiotensin III or angiotensin -(1-7) rather than by angiotensin II. The applicant proposes to receive in depth training in molecular biology, and in the process to undertake the molecular cloning and expression of aminopeptidase A, the enzyme catalyzing the formation of angiotensin III. The regulation of aminopeptidase A and of prolyl endopeptidase the enzyme catalyzing the formation of angiotensin-(1-7) will be studied in cell cultures. Our work on the development of proteinase inhibitors, with particular emphasis on inhibitors of MPC and metalloproteinases will be continued. Such compounds may represent a new generation of psychotherapeutic agents.
