Enhanced antimicrobial activity of engineered human lysozyme.
Enhanced antimicrobial activity of engineered human lysozyme.
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DOI：
10.1021/cb1001119
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发表时间：
2010-09-17
影响因子：
4
通讯作者：
Griswold, Karl E.
中科院分区：
文献类型：
--
作者：
Scanlon, Thomas C.;Teneback, Charlotte C.;Gill, Avinash;Bement, Jenna L.;Weiner, Joshua A.;Lamppa, John W.;Leclair, Laurie W.;Griswold, Karl E.
溶菌酶含有不成比例的大比例阳离子残基，因此被吸引到带负电的细菌靶标表面。重要的是，这种保守的生物物理特性可能会在急性和慢性感染期间抑制溶菌酶的抗菌功能。急性肺部铜绿假单胞菌感染的小鼠模型显示，阴离子生物聚合物在感染的肺中积累到高浓度，这些物种的存在与内源性溶菌酶活性降低有关。利用蛋白质工程技术对人溶菌酶(HLYS)的静电势进行改造，以开发专门用于呼吸道抗菌药物的抗菌酶。设计了一种新颖的高通量筛选方法，对电荷工程hLYS蛋白的组合文库进行了功能查询，并对具有更高杀菌活性的突变体进行了详细的分离和鉴定。这些研究揭示了聚阴离子抑制溶菌酶功能的一般机制，并首次直接证明了在存在与疾病相关的生物聚合物的情况下，降低hLYS的网络阳离子特性可以提高其抗菌活性。除了避免静电隔离外，至少有一种电荷工程变体在没有抑制性生物聚合物的情况下也能更快地杀灭细菌；这一观察结果支持了一种新的假说，即调节肽聚糖水解酶的细胞亲和力可能是改善细菌杀灭动力学的一般策略。
Lysozymes contain a disproportionately large fraction of cationic residues, and are thereby attracted towards the negatively charged surface of bacterial targets. Importantly, this conserved biophysical property may inhibit lysozyme antibacterial function during acute and chronic infections. A mouse model of acute pulmonary Pseudomonas aeruginosa infection demonstrated that anionic biopolymers accumulate to high concentrations in the infected lung, and the presence of these species correlates with decreased endogenous lysozyme activity. To develop antibacterial enzymes designed specifically to be used as antimicrobial agents in the infected airway, the electrostatic potential of human lysozyme (hLYS) was remodeled by protein engineering. A novel, high throughput screen was implemented to functionally interrogate combinatorial libraries of charge engineered hLYS proteins, and variants with improved bactericidal activity were isolated and characterized in detail. These studies illustrate a general mechanism by which polyanions inhibit lysozyme function, and they are the first direct demonstration that decreasing hLYS's net cationic character improves its antibacterial activity in the presence of disease-associated biopolymers. In addition to avoiding electrostatic sequestration, at least one charge engineered variant also kills bacteria more rapidly in the absence of inhibitory biopolymers; this observation supports a novel hypothesis that tuning the cellular affinity of peptidoglycan hydrolases may be a general strategy for improving kinetics of bacterial killing.
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DOI：
10.1073/pnas.0705898104
发表时间：
2007-10-09
影响因子：
11.1
作者：
Sanders, Lori K.;Xian, Wujing;Wong, Gerard C. L.
通讯作者：
Wong, Gerard C. L.
影响因子：
2.9
作者：
KNUTSON, CA;JEANES, A
通讯作者：
JEANES, A
影响因子：
14.9
作者：
Landau M;Mayrose I;Rosenberg Y;Glaser F;Martz E;Pupko T;Ben-Tal N
通讯作者：
Ben-Tal N
影响因子：
4.4
作者：
Nash, James A.;Ballard, Tiffany Nicole S.;Akinbi, Henry T.
通讯作者：
Akinbi, Henry T.
影响因子：
2.9
作者：
Krieger, E;Koraimann, G;Vriend, G
通讯作者：
Vriend, G