The objective of the proposed research is to pursue studies on glutaric acidemia type I (GAI), a human inborn error of lysine and tryptophan metabolism which causes degeneration of the basal ganglia and a progressive movement disorder in childhood, with death usually occurring during the first decade. The condition is due to deficiency of glutaryl-CoA dehydrogenase (GCDH), an FAD-containing mitochondrial enzyme that catalyzes oxidative decarboxylation of glutaryl-CoA to crotonyl-CoA and CO2. We have been investigating GAI and GCDH for some fifteen years, and are now positioned to substantially improve our understanding of how the enzyme functions in the normal state and how function is perturbed by naturally occurring mutations to cause GAI. Specific aims for this funding period are to (a) isolate glutaryl-CoA dehydrogenase from pork and human liver, and to develop amino terminal and internal peptide sequences; (b) to develop a full length cDNA clone encoding the human enzyme, authenticating its identity by colinearity of nucleotide and amino acid sequences and by expression in cos cells, fibroblasts, or frog oocytes; and (c) to develop a structure function map of GCDH by expressing normal and mutant enzymes in E coli.