CERM and Department of Chemistry Ugo Schiff, University of Florence, Via Luigi Sacconi 6, 50019, Sesto Fiorentino, Italy.
J Comput Aided Mol Des. 2012 Apr;26(4):425-35. doi: 10.1007/s10822-012-9571-0. Epub 2012 Apr 25.
Metallo-β-lactamases (MβLs) are Zn(II)-based bacterial enzymes that hydrolyze β-lactam antibiotics, hampering their beneficial effects. In the most relevant subclass (B1), X-ray crystallography studies on the enzyme from Bacillus Cereus point to either two zinc ions in two metal sites (the so-called '3H' and 'DCH' sites) or a single Zn(II) ion in the 3H site, where the ion is coordinated by Asp120, Cys221 and His263 residues. However, spectroscopic studies on the B1 enzyme from B. Cereus in the mono-zinc form suggested the presence of the Zn(II) ion also in the DCH site, where it is bound to an aspartate, a cysteine, a histidine and a water molecule. A structural model of this enzyme in its DCH mononuclear form, so far lacking, is therefore required for inhibitor design and mechanistic studies. By using force field based and mixed quantum-classical (QM/MM) molecular dynamics (MD) simulations of the protein in aqueous solution we constructed such structural model. The geometry and the H-bond network at the catalytic site of this model, in the free form and in complex with two common β-lactam drugs, is compared with experimental and theoretical findings of CphA and the recently solved crystal structure of new B2 MβL from Serratia fonticola (Sfh-I). These are MβLs from the B2 subclass, which features an experimentally well established mono-zinc form, in which the Zn(II) is located in the DCH site. From our simulations the εεδ and δεδ protomers emerge as possible DCH mono-zinc reactive species, giving a novel contribution to the discussion on the MβL reactivity and to the drug design process.
金属β-内酰胺酶（MβLs）是基于 Zn(II) 的细菌酶，可水解β-内酰胺抗生素，从而削弱其有益作用。在最相关的亚类（B1）中，对来自蜡状芽孢杆菌的酶的 X 射线晶体学研究表明，要么在两个金属位点（所谓的“3H”和“DCH”位点）中有两个锌离子，要么在 3H 位点中有一个单 Zn(II) 离子，该离子由 Asp120、Cys221 和 His263 残基配位。然而，对来自蜡状芽孢杆菌的 B1 酶的单锌形式的光谱研究表明，DCH 位点也存在 Zn(II) 离子，它与天冬氨酸、半胱氨酸、组氨酸和水分子结合。因此，需要构建该酶的 DCH 单核形式的结构模型，用于抑制剂设计和机制研究。通过使用基于力场的和混合量子经典（QM/MM）分子动力学（MD）模拟水溶液中的蛋白质，我们构建了这种结构模型。该模型在自由形式和与两种常见的β-内酰胺药物结合时的催化位点的几何形状和氢键网络与 CphA 的实验和理论发现以及最近解决的来自粘质沙雷氏菌（Serratia fonticola）的新型 B2 MβL（Sfh-I）的晶体结构进行了比较。这些是 B2 亚类的 MβLs，其具有实验上建立良好的单锌形式，其中 Zn(II) 位于 DCH 位点。从我们的模拟中，εεδ 和δεδ 原聚体作为可能的 DCH 单锌反应性物质出现，为 MβL 反应性的讨论和药物设计过程提供了新的贡献。
