The enzyme adenylosuccinate synthetase obtained from E. coli is dimeric and binds three substrates plus a metal ion. The chemical mechanism of the function of the synthetase is approached from a number of points of view. Specific amino acid residues involved in binding and catalytic processes are examined via chemical and metagenesis approaches. These studies require considerable spectroscopic effort to localize the observed effects. Chemical and spectroscopic exchange studies will be performed to get at central questions of the timing and order of binding and catalytic steps. Ongoing crystallograpic studies of wild type and mutant enzyme as well as enzyme in the presence of inhibitors will be continued. %%% Adenylosuccinate synthetase is involved in the conversion of AMP from IMP. By virtue of its location at the beginning of a metabolic pathway, this enzyme is subject to a number of controls of its catalytic activity as part of the overall metabolic integration of the cell. The proposed research explores the complex interactions which exist between the enzyme and metabolites and inhibitors it binds. The proposed research will identify the order and tightness of binding of effectors, substrates, and products, will identify catalytically important structures in the enzyme, and will identify and characterize intermediates occurring along the reaction pathway. Studies of the three-dimensional structure of the enzyme, using X-ray crystallography, will be continued.
