Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA.
Nat Commun. 2019 Jul 26;10(1):3358. doi: 10.1038/s41467-019-11230-9.
The integration of β-barrel proteins into the bacterial outer membrane (OM) is catalysed by the β-barrel assembly machinery (BAM). The central BAM subunit (BamA) itself contains a β-barrel domain that is essential for OM protein biogenesis, but its mechanism of action is unknown. To elucidate its function, here we develop a method to trap a native Escherichia coli β-barrel protein bound stably to BamA at a late stage of assembly in vivo. Using disulfide-bond crosslinking, we find that the first β-strand of a laterally 'open' form of the BamA β-barrel forms a rigid interface with the C-terminal β-strand of the substrate. In contrast, the lipid-facing surface of the last two BamA β-strands forms weaker, conformationally heterogeneous interactions with the first β-strand of the substrate that likely represent intermediate assembly states. Based on our results, we propose that BamA promotes the membrane integration of partially folded β-barrels by a 'swing' mechanism.
β-桶蛋白整合到细菌外膜（OM）是由β-桶组装机制（BAM）催化的。中央 BAM 亚基（BamA）本身含有一个β-桶结构域，对 OM 蛋白生物发生至关重要，但它的作用机制尚不清楚。为了阐明其功能，我们在这里开发了一种方法，用于在体内组装的晚期捕获稳定结合到 BamA 的天然大肠杆菌β-桶蛋白。使用二硫键交联，我们发现 BamA β-桶的横向“打开”形式的第一个β-链与底物的 C 末端β-链形成刚性界面。相比之下，最后两个 BamA β-链的脂质面向表面与底物的第一个β-链形成较弱的、构象异质的相互作用，这可能代表中间组装状态。基于我们的结果，我们提出 BamA 通过“摆动”机制促进部分折叠的β-桶的膜整合。
