PDF(Pubmed)
Abstract：
Bacteriophage endolysins are bacteriolytic enzymes that have been explored as potential weapons to fight antibiotic-resistant bacteria. Despite several studies support the application of endolysins as enzybiotics, detailed knowledge on cellular and enzymatic factors affecting their lytic activity is still missing. The bacterial membrane proton motive force (PMF) and certain cell wall glycopolymers of Gram-positive bacteria have been implicated in some tolerance to endolysins. Here, we studied how the anti-staphylococcal endolysin Lys11, a modular enzyme with two catalytic domains (peptidase and amidase) and a cell binding domain (CBD11), responded to changes in the chemical and/or electric gradients of the PMF (ΔpH and Δψ, respectively). We show that simultaneous dissipation of both gradients enhances endolysin binding to cells and lytic activity. The collapse of ΔpH is preponderant in the stimulation of Lys11 lytic action, while the dissipation of Δψ is mainly associated with higher endolysin binding. Interestingly, this binding depends on the amidase domain. The peptidase domain is responsible for most of the Lys11 bacteriolytic activity. Wall teichoic acids (WTAs) are confirmed as major determinants of endolysin tolerance, in part by severely hindering CBD11 binding activity. In conclusion, the PMF and WTA interfere differently with the endolysin functional domains, affecting both the binding and catalytic efficiencies.
摘要：
噬菌体内溶素是细菌分解酶，已被开发为对抗抗生素抗性细菌的潜在武器。尽管有几项研究支持内溶素作为酶的应用，关于影响其裂解活性的细胞和酶因子的详细知识仍然缺失。革兰氏阳性细菌的细菌膜质子动力（PMF）和某些细胞壁的糖共聚物与对内溶素的某些耐受性有关。这里,我们研究了抗葡萄球菌内溶素Lys11，一种具有两个催化域（肽酶和酰胺酶）和一个细胞结合域（CBD11）的模块化酶，对PMF的化学和/或电梯度的变化(ΔpH和ΔΦ，分别)。我们表明，两个梯度的同时消散增强了细胞内溶素与细胞的结合和裂解活性。ΔpH的崩溃在Lys11裂解作用的刺激中占优势，而ΔΦ的消散主要与较高的内溶素结合有关。有趣的是,这种结合取决于酰胺酶结构域。肽酶结构域负责大部分Lys11细菌分解活性。壁磷壁酸（WTAs）被确认为内溶素耐受性的主要决定因素，部分严重阻碍了CBD11的结合活性。总之,PMF和WTA对细胞内溶素功能结构域的干扰不同，影响结合效率和催化效率。
