College of Chemistry, MOE Key Laboratory of Green Chemistry, Sichuan University, Chengdu, Sichuan 610064, People's Republic of China.
J Phys Chem B. 2011 Sep 1;115(34):10360-7. doi: 10.1021/jp2046504. Epub 2011 Aug 5.
The dipeptide glycyl-L-tyrosine (GY) can be either a substrate for carboxypeptidase A (CPA) or an inhibitor, depending on pH. In this work, we investigate the pH-dependent reactivity of this dipeptide in CPA-catalyzed hydrolysis using a combined quantum mechanical and molecular mechanical method. It is shown that the monoionic form of the dipeptide, prevalent at high pH, chelates the active site zinc ion, rendering the enzyme inactive. This inhibitory form is consistent with an earlier X-ray structure of the CPA-GY complex. On the other hand, the prevailing di-ionic form of the dipeptide at low pH was found to undergo hydrolysis via a nucleophilic mechanism, leading to an acyl-enzyme complex. The stability of this reaction intermediate is consistent with previous low-temperature solid-state NMR results. The calculated overall free-energy barrier of 20.1 kcal/mol is in excellent agreement with the experimental value of 19.9 kcal/mol.
二肽甘氨酰-L-酪氨酸（GY）可以是羧肽酶 A（CPA）的底物，也可以是抑制剂，这取决于 pH 值。在这项工作中，我们使用量子力学和分子力学相结合的方法研究了该二肽在 CPA 催化水解中随 pH 值变化的反应性。结果表明，二肽的单离子形式在高 pH 值下占主导地位，螯合活性位点锌离子，使酶失活。这种抑制形式与之前 CPA-GY 复合物的 X 射线结构一致。另一方面，在低 pH 值下占主导地位的二肽二价形式被发现通过亲核机制发生水解，生成酰-酶复合物。该反应中间体的稳定性与先前低温固态 NMR 结果一致。计算得到的总自由能势垒为 20.1 kcal/mol，与实验值 19.9 kcal/mol 非常吻合。
