College of Chemistry, Beijing Normal University, Beijing 100875, People's Republic of China.
J Inorg Biochem. 2010 Jan;104(1):37-46. doi: 10.1016/j.jinorgbio.2009.09.025. Epub 2009 Oct 1.
The reaction mechanism of the dinuclear zinc enzyme human renal dipeptidase is investigated using hybrid density functional theory. This enzyme catalyzes the hydrolysis of dipeptides and beta-lactam antibiotics. Two different protonation states in which the important active site residue Asp288 is either neutral or ionized were considered. In both cases, the bridging hydroxide is shown to be capable of performing the nucleophilic attack on the substrate carbonyl carbon from its bridging position, resulting in the formation of a tetrahedral intermediate. This step is followed by protonation of the dipeptide nitrogen, coupled with C-N bond cleavage. The calculations establish that both cases have quite feasible energy barriers. When the Asp288 is neutral, the hydrolytic reaction occurs with a large exothermicity. However, the reaction becomes very close to thermoneutral with an ionized Asp288. The two zinc ions are shown to play different roles in the reaction. Zn1 binds the amino group of the substrate, and Zn2 interacts with the carboxylate group of the substrate, helping in orienting it for the nucleophilic attack. In addition, Zn2 stabilizes the oxyanion of the tetrahedral intermediate, thereby facilitating the nucleophilic attack.
使用杂化密度泛函理论研究了双核锌酶人肾二肽酶的反应机理。该酶催化二肽和β-内酰胺抗生素的水解。考虑了两种不同的质子化状态，其中重要的活性位点残基 Asp288 要么呈中性，要么呈离子化。在这两种情况下，桥连氢氧根都能够从桥接位置对底物羰基碳进行亲核攻击，从而形成四面体中间体。这一步之后是二肽氮的质子化，伴随着 C-N 键的断裂。计算表明这两种情况都具有相当可行的能量障碍。当 Asp288 呈中性时，水解反应具有很大的放热性。然而，当 Asp288 离子化时，反应变得非常接近热中性。两个锌离子在反应中扮演不同的角色。Zn1 结合底物的氨基，Zn2 与底物的羧酸盐基团相互作用，帮助其定向进行亲核攻击。此外，Zn2 稳定四面体中间体的含氧阴离子，从而促进亲核攻击。
