Structural and Functional Insights Into Lysostaphin-Substrate Interaction.
Structural and Functional Insights Into Lysostaphin-Substrate Interaction.
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DOI：
10.3389/fmolb.2018.00060
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发表时间：
2018
影响因子：
5
通讯作者：
Permi P
中科院分区：
文献类型：
--
作者：
Tossavainen H;Raulinaitis V;Kauppinen L;Pentikäinen U;Maaheimo H;Permi P
来自模拟葡萄球菌的溶葡萄球菌素及其家族酶作为治疗金黄色葡萄球菌感染的下一代药物迅速获得关注。溶葡萄球菌素的特异性是通过其 C 端细胞壁靶向结构域对金黄色葡萄球菌细胞壁中五甘氨酸桥的选择性来促进的。这些交联的断裂是由其 N 末端催化结构域（一种锌依赖性内肽酶）完成的。了解影响催化效率以及相互作用强度和特异性的决定因素是所有溶葡萄球菌素家族酶应用的核心。为此，我们使用NMR、SAXS和分子动力学模拟来表征溶葡萄球菌素的结构和动力学，以解决结构域间相互作用、酶-底物相互作用以及溶液中五甘氨酸裂解的催化特性。我们的核磁共振结构证实了最近的晶体结构，但与分子动力学模拟一起，强调了包含催化位点的环的动态性质。我们没有发现域间相互作用的证据，但有趣的是，SAXS 数据描绘了两个首选构象亚群。观察到催化 H329 和 H360 结合第二个锌离子，从而降低溶葡萄球菌素五甘氨酸裂解活性。发现五甘氨酸或其赖氨酸衍生物与靶向结构域的结合具有非常低的亲和力。五甘氨酸相互作用位点位于结构域的 N 末端凹槽。值得注意的是，靶向结构域以更高的微摩尔亲和力结合肽聚糖干肽 Ala-d-γ-Glu-Lys-d-Ala-d-Ala。结合位点作图揭示了该肽结构域表面上具有不同亲和力的两个相互作用位点。
Lysostaphin from Staphylococcus simulans and its family enzymes rapidly acquire prominence as the next generation agents in treatment of S. aureus infections. The specificity of lysostaphin is promoted by its C-terminal cell wall targeting domain selectivity toward pentaglycine bridges in S. aureus cell wall. Scission of these cross-links is carried out by its N-terminal catalytic domain, a zinc-dependent endopeptidase. Understanding the determinants affecting the efficiency of catalysis and strength and specificity of interactions lies at the heart of all lysostaphin family enzyme applications. To this end, we have used NMR, SAXS and molecular dynamics simulations to characterize lysostaphin structure and dynamics, to address the inter-domain interaction, the enzyme-substrate interaction as well as the catalytic properties of pentaglycine cleavage in solution. Our NMR structure confirms the recent crystal structure, yet, together with the molecular dynamics simulations, emphasizes the dynamic nature of the loops embracing the catalytic site. We found no evidence for inter-domain interaction, but, interestingly, the SAXS data delineate two preferred conformation subpopulations. Catalytic H329 and H360 were observed to bind a second zinc ion, which reduces lysostaphin pentaglycine cleaving activity. Binding of pentaglycine or its lysine derivatives to the targeting domain was found to be of very low affinity. The pentaglycine interaction site was located to the N-terminal groove of the domain. Notably, the targeting domain binds the peptidoglycan stem peptide Ala-d-γ-Glu-Lys-d-Ala-d-Ala with a much higher, micromolar affinity. Binding site mapping reveals two interaction sites of different affinities on the surface of the domain for this peptide.
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DOI：
10.1016/j.bbamem.2014.05.031
发表时间：
2015-01
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
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通讯作者：
Singh M
影响因子：
6.1
作者：
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Svergun, DI
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发表时间：
2016-09
期刊：
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影响因子：
--
作者：
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通讯作者：
Sabała I
影响因子：
11.4
作者：
Baba, T;Schneewind, O
通讯作者：
Schneewind, O
影响因子：
3.2
作者：
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通讯作者：
Schneewind, O