Molecular basis for self-assembly of a human host-defense peptide that entraps bacterial pathogens.
Molecular basis for self-assembly of a human host-defense peptide that entraps bacterial pathogens.
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DOI：
10.1021/ja5057906
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发表时间：
2014-09-24
影响因子：
15
通讯作者：
Nolan, Elizabeth M.
中科院分区：
文献类型：
--
作者：
Chairatana, Phoom;Nolan, Elizabeth M.
人α-防御素6（HD 6）是天然免疫系统的一种富含半胱氨酸的多肽。虽然HD 6是抗微生物肽家族的成员，但其在体外表现出可忽略的抗菌活性。相反，HD 6具有独特的先天免疫机制，由此它自组装成捕获病原体的寡聚体，以防止肠道上皮的微生物入侵和随后的传播。分子水平上理解为什么HD 6的功能与其他人类防御素不同仍然不清楚。为了进一步阐明HD 6的自组装过程及其生物活性，我们开发了强大的方案，用于从大肠杆菌中的过表达获得高纯度的天然和突变体HD 6。我们结合生物物理特性与生物测定来探测HD 6的结构和功能。我们报告说，本地HD 6容易自组装成细长的纤维透射电子显微镜观察，凝集革兰氏阴性和阳性细菌，并防止人类胃肠道病原体单核细胞增生李斯特菌入侵培养的哺乳动物细胞。疏水残基（F2 A、I22 T、V25 T、F29 A）的突变扰乱自组装并导致生物活性减弱。特别是，F2 A和F29 A突变体在我们的实验条件下不形成原纤维，既不凝集细菌也不阻止L.单核细胞增多症侵袭。总之，我们的研究结果表明，疏水作用是促进HD 6自组装和先天免疫功能所必需的，并表明HD 6可以提供宿主防御肠道中的李斯特菌。这项调查提供了一个及时的描述如何在氨基酸序列的变化赋予不同的生理功能的防御素家族的成员。
Human α-defensin 6 (HD6) is a 32-aa cysteine-rich peptide of the innate immune system. Although HD6 is a member of an antimicrobial peptide family, it exhibits negligible antibacterial activity in vitro. Rather, HD6 possesses a unique innate immune mechanism whereby it self-assembles into oligomers that capture pathogens to prevent microbial invasion of the intestinal epithelium and subsequent dissemination. Molecular-level understanding for why HD6 functions differently from other human defensins remains unclear. To further elucidate the HD6 self-assembly process and its biological activity, we developed robust protocols for obtaining native and mutant HD6 in high purity from overexpression in Escherichia coli. We combined biophysical characterization with biological assays to probe HD6 structure and function. We report that native HD6 readily self-assembles into elongated fibrils observable by transmission electron microscopy, agglutinates both Gram-negative and -positive bacteria, and prevents the human gastrointestinal pathogen Listeria monocytogenes from invading cultured mammalian cells. Mutation of hydrophobic residues (F2A, I22T, V25T, F29A) perturbs self-assembly and results in attenuated biological activity. In particular, the F2A and F29A mutants do not form fibrils under our experimental conditions and neither agglutinate bacteria nor prevent L. monocytogenes invasion. In total, our results demonstrate that the hydrophobic effect is essential for promoting HD6 self-assembly and innate immune function, and indicate that HD6 may provide host defense against Listeria in the gut. This investigation provides a timely description of how variations in amino acid sequence confer diverse physiological functions to members of the defensin family.
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影响因子：
3.8
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