Studies on the Mechanism and Control of Enzyme Action

从大肠杆菌中获得的腺苷琥珀酸合成酶是二聚体，与三种底物和一种金属离子结合。从多个角度探讨了合成酶作用的化学机制。参与结合和催化过程的特定氨基酸残基通过化学和代谢方法进行检测。这些研究需要相当大的光谱工作来定位观察到的效应。将进行化学和光谱交换研究，以解决结合和催化步骤的时间和顺序等核心问题。正在进行的野生型和突变型酶以及存在抑制剂的酶的结晶学研究将继续进行。腺苷琥珀酸合成酶参与了AMP从IMP转化为AMP的过程。由于它位于代谢途径的起点，这种酶作为细胞整体代谢整合的一部分，其催化活性受到许多控制。这项拟议的研究探索了酶与其结合的代谢物和抑制剂之间存在的复杂相互作用。这项拟议的研究将确定效应器、底物和产物结合的有序性和紧密性，将确定酶中具有催化作用的重要结构，并将确定和表征沿反应途径发生的中间产物。将继续使用X射线结晶学对该酶的三维结构进行研究。

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.