structural characterization of bacterial secretion channels

细菌分泌通道的结构特征

• 批准号: 8741419
• 负责人: Susan Buchanan
• 金额: $ 82.31万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8741419
• 关键词: Adopted; Bacteria; Biogenesis; C-terminal; Cell Survival; Chloroplasts; Complex; Cytoplasm; Evolution; Future; Gram-Negative Bacteria; Hemophilus ducreyi; Lateral; Lipid Bilayers; Lipoproteins; Membrane; Membrane Proteins; Mitochondria; Molecular Chaperones; Molecular Conformation; N-terminal; Names; Nature; Neisseria gonorrhoeae; Organelles; Peptidoglycan; Process; Protein translocation; Proteins; Role; Signal Transduction; Structure; Surface; Work; beta barrel; insight; interest; membrane biogenesis; periplasm; research study; simulation

Gram-negative bacteria, mitochondria, and chloroplasts contain an inner and outer membrane. The outer membrane contains a host of beta-barrel proteins commonly called outer membrane proteins (OMPs), which serve essential functions in cargo transport and signaling and are also vital for membrane biogenesis. In Gram-negative bacteria, it is known that OMPs are synthesized in the cytoplasm and then transported across the inner membrane into the periplasm via a Sec translocon. Once in the periplasm, chaperones guide the nascent OMPs across the periplasm and peptidoglycan to the inner surface of the outer membrane. Here, the nascent OMPs are recognized by a complex known as the beta-barrel assembly machinery (BAM) complex which folds and inserts the new OMPs into the outer membrane. Exactly how the BAM complex is able to accomplish its function remains unknown. However, we do know that the BAM complex consists of five components named BamA (an OMP itself) and BamB, BamC, BamD, and BamE, which are all accessory lipoproteins. Studies have shown that BamA and BamD are absolutely essential for cell viability and OMP biogenesis. Similar mechanisms for OMP biogenesis exist for both mitochondria and chloroplasts, further evidence of the evolution of these organelles. Recently, we solved the structure of BamB, while other groups solved BamC, BamD, BamE and a large portion of the periplasmic domain of BamA. Together these structures provided insight into how the BAM complex may recognize nascent OMPs. However, even with these structures being known, the mechanism for how the BAM complex recognizes, folds, and inserts nascent OMPs into the outer membrane remains elusive. To understand the mechanism of the BAM complex, we have determined crystal structures of the core membrane component called BamA, a β-barrel membrane protein itself, from two different species (Neisseria gonorrhoeae and Haemophilus ducreyi). The structure of BamA contains a large N-terminal periplasmic domain and a C-terminal 16-stranded β-barrel domain. The periplasmic domain was found in two different conformations representing open and closed states, which may serve as a gating mechanism to allow substrate access to the internal barrel cavity. Interestingly, the closed state was accompanied by a significant destabilization of the terminal strand, which was found tucked inside the barrel domain. MD simulations revealed that BamA could destabilize the local membrane along the terminal strand, thinning the membrane by as much as 16 . In addition, these MD simulations also revealed that the barrel domain of BamA may undergo a lateral opening to create a portal from the periplasm directly into the outer membrane. This work is in press at Nature. Future experiments will investigate the roles of the 4 BAM lipoproteins and how they assemble and function together.

革兰氏阴性菌、线粒体和叶绿体含有内膜和外膜。 外膜含有一系列通常称为外膜蛋白（OMP）的β桶蛋白，它们在货物运输和信号传导中发挥重要作用，对膜生物发生也至关重要。在革兰氏阴性细菌中，已知OMP在细胞质中合成，然后经由Sec易位子穿过内膜转运到周质中。一旦在周质中，伴侣蛋白引导新生的OMP穿过周质和肽聚糖到达外膜的内表面。在这里，新生的OMPs被一种称为β桶组装机制（BAM）复合物的复合物识别，该复合物折叠并将新的OMPs插入外膜中。BAM复合体究竟是如何实现其功能的仍然是未知的。然而，我们知道BAM复合物由五种成分组成，分别为BamA（本身是一种OMP）和BamB、BamC、BamD和BamE，它们都是辅助脂蛋白。研究表明，BamA和BamD对细胞活力和OMP生物合成是绝对必要的。线粒体和叶绿体都存在类似的OMP生物发生机制，这进一步证明了这些细胞器的进化。最近，我们解决了BamB的结构，而其他小组解决了BamC，BamD，BamE和BamA的大部分周质结构域。这些结构一起提供了对BAM复合物如何识别新生OMP的深入了解。然而，即使这些结构是已知的，BAM复合物如何识别，折叠，并插入新生的外膜外蛋白的机制仍然难以捉摸。 为了理解BAM复合物的机制，我们已经确定了来自两个不同物种（淋病奈瑟菌和杜克雷嗜血杆菌）的称为BamA的核心膜组分（桶膜蛋白本身）的晶体结构。BamA的结构包含一个大的N-末端周质结构域和一个C-末端16链桶结构域。周质域被发现在两个不同的构象代表开放和封闭的状态，这可能是一个门控机制，允许基板进入内部桶腔。有趣的是，闭合状态伴随着末端链的显著不稳定，这被发现隐藏在桶结构域内。分子动力学模拟显示，BamA可以破坏局部膜沿着末端链，使膜变薄多达16。此外，这些分子动力学模拟还揭示了BamA的桶结构域可能经历了一个侧向开口，以创建一个从周质直接进入外膜的门户。这项工作正在《自然》杂志上出版。未来的实验将研究4种BAM脂蛋白的作用以及它们如何组装和共同发挥作用。