Acceleration of Enzymatic Reaction of Trypsin through the Formation of Water-Soluble Complexes with Poly(ethylene glycol)-block-Poly(α,β-aspartic acid)
Acceleration of Enzymatic Reaction of Trypsin through the Formation of Water-Soluble Complexes with Poly(ethylene glycol)-block-Poly(α,β-aspartic acid)
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DOI：
10.1021/bm049198w
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发表时间：
2005-02
影响因子：
6.2
通讯作者：
Akifumi Kawamura;Yuriko Yoshioka;A. Harada;K. Kono
中科院分区：
文献类型：
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作者：
Akifumi Kawamura;Yuriko Yoshioka;A. Harada;K. Kono
以L-赖氨酸对硝基苯胺为底物，采用比色法测定了牛胰胰蛋白酶与聚乙二醇-聚α，β-天冬氨酸（PEG-PAA）形成的水溶性复合物的酰胺酶活性。PEG-PAA与胰蛋白酶的络合作用加速了胰蛋白酶的酶促反应。通过测定胰蛋白酶的酶促反应动力学参数，证实复合胰蛋白酶的催化速率常数比天然胰蛋白酶高15倍。从初始反应速率的pH依赖性的评价，它表明，这种加速是由稳定的His残基的咪唑鎓离子的催化位点，Asp-His-Ser三联体，胰蛋白酶由于天冬氨酸单位的PEG-PAA诱导。氢键Asp-His对是丝氨酸蛋白酶和脱嘌呤核酸内切酶等几种关键酶促反应的关键组成部分，预计在其他条件下可能会发生催化反应的加速。
The amidase activity of bovine pancreas trypsin in water-soluble complexes with poly(ethylene glycol)-block-poly(α,β-aspartic acid) (PEG-PAA) was evaluated by a colorimetric assay using l-lysine p-nitroanilide as a substrate. The enzymatic reaction of trypsin was accelerated through the complexation with PEG-PAA. By determining the kinetic parameters of the enzymatic reaction of trypsin, it was confirmed that the catalytic rate constant of the complexed trypsin was 15 times higher than that of the native trypsin. From the evaluation of pH dependence of initial reaction rate, it was indicated that this acceleration was induced by a stabilization of the imidazolium ion of the His residue in the catalytic site, the Asp-His-Ser triad, of trypsin due to the Asp units of PEG-PAA. The hydrogen bonded Asp-His pairs are critical constituents in several key enzymatic reactions including serine protease and apurinic endonucleases, and it was expected that the acceleration of the catalytic reaction might occur for ot...