Functions of the BamBCDE Lipoproteins Revealed by Bypass Mutations in BamA
Functions of the BamBCDE Lipoproteins Revealed by Bypass Mutations in BamA
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DOI：
10.1128/jb.00401-20
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发表时间：
2020-11-01
影响因子：
3.2
通讯作者：
Silhavy, Thomas J.
中科院分区：
文献类型：
--
作者：
Hart, Elizabeth M.;Silhavy, Thomas J.
异五聚体β-桶组装机（BAM复合物）负责将多种β-桶外膜蛋白（OMP）折叠并插入革兰氏阴性菌的外膜（OM）中。BAM复合物包含两种必需蛋白质，β-桶OMP BamA和脂蛋白BamD，而辅助脂蛋白BamBCE单独为非必需的。在这里，我们识别并表征了三种bamA突变，即位置470处的E至K变化（bamA（E470 K））、位置496处的A至P变化（bamA（A496 P））和位置499处的A至S变化（bamA（A499 S）），它们抑制了原本致命的Delta bamD、Delta bamB Delta bamC Delta bamE和Delta bamC Delta bamD Delta bamE突变。缺乏BAM复合脂蛋白的不同组合的细胞的活力提供了检查OMP组装中单个蛋白质的作用的机会。结果表明，在野生型细胞中，BamBCE共享冗余功能;这些脂蛋白中的至少一种必须存在以允许BamD与BamA有效地协调。除了BamA调节外，BamD还具有一个额外的基本功能，该功能与BamB的第二功能是多余的。值得注意的是，bamA（E470 K）抑制这两种蛋白，允许构建仅由BamA（E470 K）组成的BAM复合物，其能够在不存在BamBCDE的情况下组装OMP。这项工作表明，BAM复合物脂蛋白不参与催化折叠的OMP基板，而是功能，以提高组装过程的效率，通过协调和调节组装的不同OMP substrate.IMPORTANCE的折叠和插入的β-桶外膜蛋白（OMP）是保守的过程中的线粒体，叶绿体，和革兰氏阴性菌。在革兰氏阴性菌中，外膜蛋白通过异五聚体β-桶组装机（BAM复合物）组装到外膜（OM）中。在这项研究中，我们探讨了个别BAM蛋白的功能，以及它们如何协调组装的一个不同的家庭外膜蛋白。此外，我们确定了一个增益功能bamA突变体能够组装OMPs独立于所有其他四个BAM蛋白。这项工作推进了我们对OMP组装的理解，并揭示了这一过程在革兰氏阴性菌中的独特性。
The heteropentomeric beta-barrel assembly machine (BAM complex) is responsible for folding and inserting a diverse array of beta-barrel outer membrane proteins (OMPs) into the outer membrane (OM) of Gram-negative bacteria. The BAM complex contains two essential proteins, the beta-barrel OMP BamA and a lipoprotein BamD, whereas the auxiliary lipoproteins BamBCE are individually nonessential. Here, we identify and characterize three bamA mutations, the E-to-K change at position 470 (bamA(E470K) ), the A-to-P change at position 496 (bamA(A496P)), and the A-to-S change at position 499 (bamA(A499S)), that suppress the otherwise lethal Delta bamD, Delta bamB Delta bamC Delta bamE, and Delta bamC Delta bamD Delta bamE mutations. The viability of cells lacking different combinations of BAM complex lipoproteins provides the opportunity to examine the role of the individual proteins in OMP assembly. Results show that, in wild-type cells, BamBCE share a redundant function; at least one of these lipoproteins must be present to allow BamD to coordinate productively with BamA. Besides BamA regulation, BamD shares an additional essential function that is redundant with a second function of BamB. Remarkably, bamA(E470K) suppresses both, allowing the construction of a BAM complex composed solely of BamA(E470K) that is able to assemble OMPs in the absence of BamBCDE. This work demonstrates that the BAM complex lipoproteins do not participate in the catalytic folding of OMP substrates but rather function to increase the efficiency of the assembly process by coordinating and regulating the assembly of diverse OMP substrates.IMPORTANCE The folding and insertion of beta-barrel outer membrane proteins (OMPs) are conserved processes in mitochondria, chloroplasts, and Gram-negative bacteria. In Gram-negative bacteria, OMPs are assembled into the outer membrane (OM) by the heteropentomeric beta-barrel assembly machine (BAM complex). In this study, we probe the function of the individual BAM proteins and how they coordinate assembly of a diverse family of OMPs. Furthermore, we identify a gain-offunction bamA mutant capable of assembling OMPs independently of all four other BAM proteins. This work advances our understanding of OMP assembly and sheds light on how this process is distinct in Gram-negative bacteria.