ES complexes of Aeromonas aminopeptidase: direct observation by stopped-flow fluorescence.
ES complexes of Aeromonas aminopeptidase: direct observation by stopped-flow fluorescence.
复制标题
气单胞菌氨肽酶的 ES 复合物：通过停流荧光直接观察。
DOI：
10.1016/0006-291x(83)91391-8
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发表时间：
1983
影响因子：
3.1
通讯作者：
J. M. Prescott
中科院分区：
文献类型：
--
作者：
D. Auld;J. M. Prescott
气单胞菌氨肽酶的中间体是通过荧光监测所产生的无辐射能量转移（RET）之间的酶和结合底物的丹磺酰基基团。在结合底物酶时，色氨酸荧光被淬灭，底物丹磺酰荧光增强。这些过程在Leu-Ala键水解和Ala-DED从酶释放后逆转。在pH 7.5和20° C下，停流RET动力学分析得出的k cat= 36 sec− 1和Km = 3.7 μM。这些值代表气单胞菌氨肽酶任何底物的最高k cat Km比，1× 10 7 M− 1 sec− 1。该肽的优良结合特性允许即使在10− 7 M的酶浓度下也能直接观察ES复合物。
Intermediates of Aeromonas aminopeptidase are monitored through fluorescence generated by radiationless energy transfer (RET) between enzyme tryptophans and the dansyl group of the bound substrate. Upon binding of the substrate enzyme tryptophan fluorescence is quenched and substrate dansyl fluorescence enhanced. These processes are reversed upon hydrolysis of the Leu-Ala bond and release of Ala-DED from the enzyme. Stopped-flow RET kinetic analysis yields values of k cat= 36 sec− 1 and K m= 3.7 μM at pH 7.5 and 20° C. These values represent the highest k cat K m ratio, 1× 10 7 M− 1 sec− 1, of any substrate for Aeromonas aminopeptidase. The excellent binding properties of the peptide permit direct visualization of ES complexes even at enzyme concentrations of 10− 7 M.
金属结合化学计量和金属离子调节猪肾亮氨酸氨肽酶活性的机制。
影响因子：
2.9
作者：
VanWart,HE;Lin,SH
通讯作者：
Lin,SH