Anton D L, Kutny R
Central Research and Development Department, Experimental Station, E. I. du Pont de Nemours and Company, Wilmington, Delaware 19898.
Biochemistry. 1987 Oct 6;26(20):6444-7. doi: 10.1021/bi00394a022.
S-Adenosylmethionine decarboxylase, a pyruvoyl-containing decarboxylase, is inactivated in a time-dependent process under turnover conditions. The inactivation is dependent on the presence of both substrate and Mg2+, which is also required for enzyme activity. The rate of inactivation is dependent on the concentration of substrate and appears to be saturable. Inactivation by [methionyl-3,4-14C]-adenosylmethionine results in stoichiometric labeling of the protein. In contrast, when either S-[methyl-3H]adenosylmethionine or [8-14C]adenosylmethionine is used, there is virtually no incorporation of radioactivity. Automated Edman degradation of the alpha (pyruvoyl-containing) subunit reveals that substrate inactivation results in the conversion of the pyruvoyl group to an alanyl residue. These data suggest a mechanism of inactivation which involves the transamination of the nascent product to the pyruvoyl group, followed by the elimination of methylthioadenosine and the generation of a 2-propenal equivalent which could undergo a Michael addition to the enzyme. This is the first evidence for a transamination mechanism for substrate inactivation of a pyruvoyl enzyme.
S-腺苷甲硫氨酸脱羧酶是一种含丙酮酸基的脱羧酶，在周转条件下会以时间依赖性过程失活。这种失活依赖于底物和Mg2+的存在，而Mg2+也是酶活性所必需的。失活速率取决于底物浓度，且似乎具有饱和性。用[甲硫氨酰-3,4-14C]-腺苷甲硫氨酸进行失活处理会导致蛋白质的化学计量标记。相比之下，当使用S-[甲基-3H]腺苷甲硫氨酸或[8-14C]腺苷甲硫氨酸时，几乎没有放射性掺入。对α（含丙酮酸基）亚基进行自动埃德曼降解显示，底物失活导致丙酮酸基团转化为丙氨酰残基。这些数据提示了一种失活机制，该机制涉及新生产物与丙酮酸基团的转氨作用，随后消除甲硫基腺苷并生成一个2-丙烯醛等价物，该等价物可能会对酶进行迈克尔加成。这是关于丙酮酸基酶底物失活的转氨机制的首个证据。
