INHIBITION OF PAPAIN BY NITRILES - MECHANISTIC STUDIES USING NMR AND KINETIC MEASUREMENTS
INHIBITION OF PAPAIN BY NITRILES - MECHANISTIC STUDIES USING NMR AND KINETIC MEASUREMENTS
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DOI：
10.1016/0003-9861(87)90068-3
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发表时间：
1987-02-01
影响因子：
3.9
通讯作者：
ABELES, RH
中科院分区：
文献类型：
--
作者：
LIANG, TC;ABELES, RH
N-（N-乙酰基-1-苯丙氨酰）氨基乙腈是木瓜蛋白酶的抑制剂。用13 C NMR光谱，我们已经表明，可逆的共价加合物与木瓜蛋白酶形成。共价加合物形成的可逆性与NMR饱和转移技术使用DANTE脉冲选择性激发证明。此外，用醛抑制剂置换共价加合物以再生硝基化合物。未观察到腈水解。共价加合物很可能是基本硫醇和硝基之间形成的硫代亚胺酯。对几种对硝基苯胺底物及其相应的腈类抑制剂进行了研究。Ki与kcat/Km之间存在相关性。这一发现连同Ki的pH依赖性与Kcat/Km平行的事实表明腈和木瓜蛋白酶的相互作用具有相当大的过渡态特征。相比之下，腈显示为α-氨基甲酸酯的无效抑制剂。糜蛋白酶
N-(N-acetyl-1-phenylalanyl)aminoacetronitrile is an inhibitor of papain. With 13C NMR spectroscopy we have shown that a reversible covalent adduct is formed with papain. The reversible nature of the covalent-adduct formation was demonstrated with NMR saturation-transfer technique using a DANTE pulse for selective excitation. In addition the covalent adduct was displaced with an aldehyde inhibitor to regenerate the nitrole compound. No hydrolysis of the nitrile was observed. The covalent adduct is mostly likely a thioimidate formed between the essential thiol and the nitrole. Several p-nitroanilide substrates and their corresponding nitrile inhibitors were examined. A correlation between Ki and kcat/Km was observed. This finding together with the fact that the pH dependence of Ki parallels that kcat/Km suggests that interaction of nitriles and papain has considerable transition-state character. In contrast, a nitrile was shown to be an ineffective inhibitor of .alpha.-chymotrypsin.