Department of Microbiology, Cornell University, Ithaca, NY, 14853, USA.
Robert Frederick Smith School of Chemical and Biomolecular Engineering, Cornell University, 120 Olin Hall, Ithaca, NY, 14853, USA.
Sci Rep. 2022 Sep 1;12(1):14862. doi: 10.1038/s41598-022-18958-3.
The twin-arginine translocation (Tat) pathway involves an inbuilt quality control (QC) system that synchronizes the proofreading of substrate protein folding with lipid bilayer transport. However, the molecular details of this QC mechanism remain poorly understood. Here, we hypothesized that the conformational state of Tat substrates is directly sensed by the TatB component of the bacterial Tat translocase. In support of this hypothesis, several TatB variants were observed to form functional translocases in vivo that had compromised QC activity as evidenced by the uncharacteristic export of several misfolded protein substrates. These variants each possessed cytoplasmic membrane-extrinsic domains that were either truncated or mutated in the vicinity of a conserved, highly flexible α-helical domain. In vitro folding experiments revealed that the TatB membrane-extrinsic domain behaved like a general molecular chaperone, transiently binding to highly structured, partially unfolded intermediates of a model protein, citrate synthase, in a manner that prevented its irreversible aggregation and stabilized the active species. Collectively, these results suggest that the Tat translocase may use chaperone-like client recognition to monitor the conformational status of its substrates.
双精氨酸转运（Tat）途径涉及一个内置的质量控制系统，该系统可协调底物蛋白折叠的校对与脂双层转运。然而，这个 QC 机制的分子细节仍知之甚少。在这里，我们假设 Tat 底物的构象状态可被细菌 Tat 转运体的 TatB 组分直接感知。支持这一假说的是，观察到几种 TatB 变体在体内形成了功能性转运体，但 QC 活性受损，这表现在几种错误折叠的蛋白质底物的异常输出。这些变体都具有细胞质膜外在域，这些域要么在保守的、高度灵活的α螺旋域附近被截断，要么被突变。体外折叠实验表明，TatB 膜外在域的行为类似于一般的分子伴侣，以阻止其不可逆聚集并稳定活性物质的方式，短暂地结合到模型蛋白柠檬酸合酶的高度结构化、部分展开的中间体上。总的来说，这些结果表明 Tat 转运体可能使用类似伴侣的客户识别来监测其底物的构象状态。
