Stewart Alesha C, Bethel Christopher R, VanPelt Jamie, Bergstrom Alex, Cheng Zishuo, Miller Callie G, Williams Cameron, Poth Robert, Morris Matthew, Lahey Olivia, Nix Jay C, Tierney David L, Page Richard C, Crowder Michael W, Bonomo Robert A, Fast Walter
Division of Chemical Biology and Medicinal Chemistry, College of Pharmacy, and the LaMontagne Center for Infectious Disease, University of Texas , Austin, Texas 78712, United States.
Research Services, Louis Stokes Cleveland Department of Veterans Affairs Medical Center , Cleveland, Ohio 44106, United States.
ACS Infect Dis. 2017 Dec 8;3(12):927-940. doi: 10.1021/acsinfecdis.7b00128. Epub 2017 Oct 11.
Use and misuse of antibiotics have driven the evolution of serine β-lactamases to better recognize new generations of β-lactam drugs, but the selective pressures driving evolution of metallo-β-lactamases are less clear. Here, we present evidence that New Delhi metallo-β-lactamase (NDM) is evolving to overcome the selective pressure of zinc(II) scarcity. Studies of NDM-1, NDM-4 (M154L), and NDM-12 (M154L, G222D) demonstrate that the point mutant M154L, contained in 50% of clinical NDM variants, selectively enhances resistance to the penam ampicillin at low zinc(II) concentrations relevant to infection sites. Each of the clinical variants is shown to be progressively more thermostable and to bind zinc(II) more tightly than NDM-1, but a selective enhancement of penam turnover at low zinc(II) concentrations indicates that most of the improvement derives from catalysis rather than stability. X-ray crystallography of NDM-4 and NDM-12, as well as bioinorganic spectroscopy of dizinc(II), zinc(II)/cobalt(II), and dicobalt(II) metalloforms probe the mechanism of enhanced resistance and reveal perturbations of the dinuclear metal cluster that underlie improved catalysis. These studies support the proposal that zinc(II) scarcity, rather than changes in antibiotic structure, is driving the evolution of new NDM variants in clinical settings.
抗生素的使用和滥用促使丝氨酸β-内酰胺酶不断进化，以更好地识别新一代β-内酰胺类药物，但驱动金属β-内酰胺酶进化的选择压力尚不清楚。在此，我们提供证据表明，新德里金属β-内酰胺酶（NDM）正在进化以克服锌（II）缺乏的选择压力。对NDM-1、NDM-4（M154L）和NDM-12（M154L，G222D）的研究表明，临床NDM变体中有50%含有点突变M154L，在与感染部位相关的低锌（II）浓度下，该突变选择性地增强了对青霉烯类氨苄西林的耐药性。结果显示，每种临床变体都比NDM-1具有更高的热稳定性，并且与锌（II）的结合更紧密，但在低锌（II）浓度下对青霉烯类周转率的选择性增强表明，大部分改善源自催化作用而非稳定性。NDM-4和NDM-12的X射线晶体学研究，以及二价锌、锌（II）/钴（II）和二价钴金属形式的生物无机光谱研究，探究了耐药性增强的机制，并揭示了双核金属簇的扰动是催化作用改善的基础。这些研究支持了这样一种观点，即在临床环境中，是锌（II）缺乏而非抗生素结构的变化推动了新的NDM变体的进化。
