通过\(^{79}Br\)和\(^{81}Br\)脉冲核磁共振研究4-溴汞基肉桂酸与α-糜蛋白酶的相互作用。

Study of the interaction of 4-bromomercuriocinnamic acid with alpha-chymotrypsin by 79 Br and 81Br pulsed nuclear-magnetic resonance.

作者信息

Garnett M W, Halstead T K, Hoare D G

出版信息

Eur J Biochem. 1976 Jun 15;66(1):85-93. doi: 10.1111/j.1432-1033.1976.tb10428.x.

DOI:10.1111/j.1432-1033.1976.tb10428.x

Abstract

Br- has been used as a nuclear magnetic resonance (NMR) probe to study the reversible association of alpha-chymotrypsin and an Hg-labelled substrate (4-bromomercuriocinnamic acid, BrHgCin) which rapidly exchanges Br-. T1 was measured for 79Br and 81Br, using a pulse spectrometer. Values of the parameters that determine T1, Obs in aqueous solutions of KBr (pH=5.5) containing alpha-chymotrypsin and BrHgCin are reported. It is found that the rate of Br exchange is diffusion-limited and faster than the rate of reorientation of the BrHgCin-alpha-chymotrypsin complex. The rate constant for the formation of the covalent BrHgCin-alpha-chymotrypsin complex determined by this technique agrees well with previously published data. The rapid rate of Br exchange with the complex, however, is incompatible with the side chain of BrHgCin being entirely buried in a nonpolar pocket on the enzyme but compatible with the side chain being exposed to the solution. The contribution to the NMR signal from the non-covalent complex is negligible.

摘要

溴离子（Br⁻）已被用作核磁共振（NMR）探针，以研究α-胰凝乳蛋白酶与汞标记底物（4-溴汞基肉桂酸，BrHgCin）的可逆缔合，该底物能快速交换Br⁻。使用脉冲光谱仪测量了⁷⁹Br和⁸¹Br的T1值。报告了在含有α-胰凝乳蛋白酶和BrHgCin的KBr水溶液（pH = 5.5）中，决定T1观测值的参数值。研究发现，Br交换速率受扩散限制，且比BrHgCin-α-胰凝乳蛋白酶复合物的重排速率快。用该技术测定的共价BrHgCin-α-胰凝乳蛋白酶复合物形成的速率常数与先前发表的数据吻合良好。然而，Br与复合物的快速交换速率与BrHgCin的侧链完全埋在酶的非极性口袋中不相符，但与侧链暴露于溶液中的情况相符。非共价复合物对NMR信号的贡献可忽略不计。