The polymyxin colistin is an agent of last resort for treatment of severe infections by multiresistant Gram-negative opportunistic bacteria. Colistin resistance arises through covalent modification of lipid A disaccharide by phosphoethanolamine (PEtN) transferases, preventing colistin interaction. The Mobile Colistin Resistance (MCR) PEtN transferase is a plasmid-borne enzyme that is the major cause of colistin resistance in Escherichia coli, the most important antimicrobial resistant bacterial pathogen worldwide. Bacterial PEtN transferases like MCR comprise periplasmic catalytic and integral membrane domains, with mechanistic understanding largely based on studies of the former with limited information on the full-length enzyme. Previous investigations of a Neisseria meningitidis PEtN transferase identified that the catalytic domain can effectively dissociate from the transmembrane component and instead make extensive contacts with the membrane surface. Here we report extended molecular dynamics simulations of a model of full-length MCR-1, in a representative membrane comprising 80% of a PEtN donor substrate palmitoyloleoyl phosphoethanolamine (POPE) that explore the dynamic behavior of the enzyme and the impact upon it of zinc stoichiometry and of PEtN addition to the Thr285 acceptor residue. The results identify only limited movement of the two domains relative to one another, and that POPE can bind the likely “resting” state of the enzyme (mono-zinc with unmodified Thr285) in an orientation compatible with PEtN transfer to Thr285. Our data suggest domain motions in bacterial PEtN transferases to be condition-dependent and support a proposed “ping - pong” reaction mechanism with the mono-zinc enzyme competent to undertake the first stage.

中文翻译：

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通过膜环境中的扩展分子动力学模拟研究多粘菌素耐药决定因素 MCR-1 的动态行为和底物相互作用

多粘菌素粘菌素是治疗由多重耐药革兰氏阴性机会性细菌引起的严重感染的最后手段。粘菌素耐药性是通过磷酸乙醇胺 (PEtN) 转移酶对脂质 A 二糖进行共价修饰而产生的，从而防止粘菌素相互作用。移动粘菌素耐药性 (MCR) PEtN 转移酶是一种质粒携带的酶，是大肠杆菌（全世界最重要的抗菌药物耐药性细菌病原体）中粘菌素耐药性的主要原因。像 MCR 这样的细菌 PEtN 转移酶包含周质催化域和整合膜域，其机制的理解主要基于对前者的研究，而有关全长酶的信息有限。先前对脑膜炎奈瑟氏球菌 PEtN 转移酶的研究发现，催化结构域可以有效地从跨膜成分上解离，而是与膜表面广泛接触。在这里，我们报告了全长 MCR-1 模型的扩展分子动力学模拟，该模型在包含 80% PEtN 供体底物棕榈酰油酰磷酸乙醇胺 (POPE) 的代表性膜中进行，探索了酶的动态行为以及锌对其的影响化学计量和 PEtN 添加到 Thr285 受体残基的结果。结果表明，两个结构域相对于彼此仅进行有限的运动，并且 POPE 可以以与 PEtN 转移至 Thr285 相容的方向结合酶（具有未修饰的 Thr285 的单锌）的可能“静止”状态。我们的数据表明，细菌 PEtN 转移酶中的结构域运动是条件依赖性的，并支持所提出的“乒乓”反应机制，其中单锌酶能够进行第一阶段。