Increasing the Antimicrobial Activity of Nisin-Based Lantibiotics against Gram-Negative Pathogens
Increasing the Antimicrobial Activity of Nisin-Based Lantibiotics against Gram-Negative Pathogens
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DOI：
10.1128/aem.00052-18
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发表时间：
2018-06-01
影响因子：
4.4
通讯作者：
Kuipers, Oscar P.
中科院分区：
文献类型：
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作者：
Li, Qian;Montalban-Lopez, Manuel;Kuipers, Oscar P.
羊毛硫抗生素是核糖体合成和后修饰的抗微生物化合物，含有羊毛硫素和甲基羊毛硫素残基。乳链菌肽是研究和使用最广泛的羊毛硫抗生素之一，已被证明对革兰氏阳性菌显示出非常有效的活性，并且很少观察到稳定的耐药性。通过与脂质II结合并在膜中形成孔，乳链菌肽可以引起细胞成分的流出并抑制细胞壁生物合成。然而，乳酸链球菌素对革兰氏阴性菌的活性远低于对革兰氏阳性菌的活性，这主要是因为脂质II位于内膜，而革兰氏阴性菌中相当不渗透的外膜阻止乳酸链球菌素到达脂质II。因此，如果能够提高乳链菌肽的外膜穿透效率，则原则上能够提高抗革兰氏阴性菌的活性。在这项工作中，从文献数据中选择了几个相对较短的具有抗革兰氏阴性菌活性的肽，以作为尾部融合到完整或截短的乳链菌肽种类的C末端。在这些中，我们发现附着于乳链菌肽的一部分的三个尾（尾2 [T2; DKYLPRPRPV]、T6 [NGVQPKY]和T8 [KIAKVALKAL]）中的一个显示出针对革兰氏阴性微生物的改善的活性。接下来，我们合理地设计和重组最有前途的融合肽。获得了几个突变体，其活性显着优于乳链菌肽对革兰氏阴性病原菌。尾16突变体2（T16 m2）构建体对几种重要的革兰氏阴性病原体（即，大肠杆菌、肺炎克雷伯氏菌、鲍曼不动杆菌、铜绿假单胞菌、产气肠杆菌）与乳链菌肽相比增加4- 12倍。本研究表明，通过对乳链菌肽的合理设计，可以选择性地显著提高其外膜透过能力，同时提高其对革兰氏阴性菌的抗菌活性。重要提示羊毛硫抗生素是一种对革兰氏阳性菌具有较高活性，但对大多数革兰氏阴性菌活性相对较差的抗菌肽。在这里，我们修改了模型lantibiotic nisin融合的部分，它的抗菌肽与已知的活性对革兰氏阴性菌。适当选择可连接至乳链菌肽（部分）的肽部分可导致其对革兰氏阴性菌的抑制活性显著增加。使用这种策略，产生了通过显示针对相关革兰氏阴性细菌物种的4至12倍高水平的活性而优于乳链菌肽的杂交种。这项研究显示了修饰的肽工程在所需方向上改变靶特异性的能力。
Lantibiotics are ribosomally synthesized and posttranslationally modified antimicrobial compounds containing lanthionine and methyl-lanthionine residues. Nisin, one of the most extensively studied and used lantibiotics, has been shown to display very potent activity against Gram-positive bacteria, and stable resistance is rarely observed. By binding to lipid II and forming pores in the membrane, nisin can cause the efflux of cellular constituents and inhibit cell wall biosynthesis. However, the activity of nisin against Gram-negative bacteria is much lower than that against Gram-positive bacteria, mainly because lipid II is located at the inner membrane, and the rather impermeable outer membrane in Gram-negative bacteria prevents nisin from reaching lipid II. Thus, if the outer membrane-traversing efficiency of nisin could be increased, the activity against Gram-negative bacteria could, in principle, be enhanced. In this work, several relatively short peptides with activity against Gram-negative bacteria were selected from literature data to be fused as tails to the C terminus of either full or truncated nisin species. Among these, we found that one of three tails (tail 2 [T2; DKYLPRPRPV], T6 [NGVQPKY], and T8 [KIAKVALKAL]) attached to a part of nisin displayed improved activity against Gram-negative microorganisms. Next, we rationally designed and reengineered the most promising fusion peptides. Several mutants whose activity significantly outperformed that of nisin against Gramnegative pathogens were obtained. The activity of the tail 16 mutant 2 (T16m2) construct against several important Gram-negative pathogens (i.e., Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter aerogenes) was increased 4- to 12-fold compared to that of nisin. This study indicates that the rational design of nisin can selectively and significantly improve its outer membrane-permeating capacity as well as its activity against Gram-negative pathogens.IMPORTANCE Lantibiotics are antimicrobial peptides that are highly active against Gram-positive bacteria but that have relatively poor activity against most Gramnegative bacteria. Here, we modified the model lantibiotic nisin by fusing parts of it to antimicrobial peptides with known activity against Gram-negative bacteria. The appropriate selection of peptidic moieties that could be attached to (parts of) nisin could lead to a significant increase in its inhibitory activity against Gram-negative bacteria. Using this strategy, hybrids that outperformed nisin by displaying 4- to 12-fold higher levels of activity against relevant Gram-negative bacterial species were produced. This study shows the power of modified peptide engineering to alter target specificity in a desired direction.