Remy M H, Bourdillon C, Thomas D
Biochim Biophys Acta. 1985 May 20;829(1):69-75. doi: 10.1016/0167-4838(85)90069-x.
The aim of this work was to study the role of hydrophobic interactions in the enzymic activity of chymotrypsin. The amino groups of chymotrypsin were chemically modified by aliphatic aldehydes of various chain lengths - acetaldehyde, butyraldehyde, hexanal - and with two aldehydes of different steric hindrance - benzaldehyde and trimethyl acetaldehyde. After a rapid study of the derivated enzymes, the hexylchymotrypsin has been chosen for its new catalytic properties: the Michaelis constant is not modified and the maximal velocity with N-glutaryl-L-phenylalanine-4-nitroaniline is increased to 164%. The increase is due to the increase of the acylation constant, k2, by 230%. The value of k3 is not modified or less modified. In the modified enzyme, 85% of free amino acids are still able to react with trinitrobenzenesulphonic acid. The optimum pH is shifted by one pH unit towards the alkaline pH. The thermodynamic study shows that the catalytic process itself is not modified. The increase in Vm could be a simple increase of k2 linked to a modification of the site or of the protein. The phenomenon described is very specific and obtained only with one modification, hexanal, and with one enzyme, alpha-chymotrypsin.
这项工作的目的是研究疏水相互作用在胰凝乳蛋白酶酶活性中的作用。胰凝乳蛋白酶的氨基用不同链长的脂肪醛——乙醛、丁醛、己醛——以及两种具有不同空间位阻的醛——苯甲醛和三甲基乙醛进行化学修饰。在对衍生酶进行快速研究后，己基胰凝乳蛋白酶因其新的催化特性被选中：米氏常数未改变，以N - 谷氨酰 - L - 苯丙氨酸 - 4 - 硝基苯胺为底物时最大反应速度提高到164%。这种提高是由于酰化常数k2增加了230%。k3的值未改变或改变较小。在修饰后的酶中，85%的游离氨基酸仍能与三硝基苯磺酸反应。最适pH向碱性pH偏移了一个pH单位。热力学研究表明催化过程本身未改变。Vm的增加可能只是与位点或蛋白质修饰相关的k2的简单增加。所描述的现象非常特殊，仅通过一种修饰（己醛）和一种酶（α - 胰凝乳蛋白酶）获得。
