The sacrificial adaptor protein Skp functions to remove stalled substrates from the β-barrel assembly machine.
The sacrificial adaptor protein Skp functions to remove stalled substrates from the β-barrel assembly machine.
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DOI：
10.1073/pnas.2114997119
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发表时间：
2022-01-04
影响因子：
11.1
通讯作者：
Silhavy TJ
中科院分区：
文献类型：
--
作者：
Combs AN;Silhavy TJ
革兰氏阴性菌的外膜（OM）作为一个强大的渗透性屏障，使细胞能够在各种恶劣的环境中生存。对于OM完整性至关重要的是β-桶外膜蛋白（OMP），其通过广泛保守的β-桶组装机（BAM）复合物组装到膜中。在这里，我们确定了特定的作用，周质分子伴侣Skp作为牺牲衔接蛋白的功能，以消除停滞的基板从BAM复合物，施加一个积极的质量控制机制，确保有效的组装新生OMPs到OM。这项工作确定了Skp/DegP功能关系的分子机制，并澄清了长期存在的矛盾，即如何在体内实现从高亲和力、长寿命的Skp-OMP复合物中释放底物。革兰氏阴性菌中完整β-桶外膜蛋白（OMP）的生物发生需要分子伴侣穿过水周质空间的转运。部分由于周质分子伴侣网络中广泛的功能冗余，分子伴侣在OMP质量控制和组装中的特定作用在很大程度上仍然是难以捉摸的。在这里，通过使用多个折叠缺陷底物故意干扰OMP组装过程，我们已经确定了周质伴侣Skp在确保β-桶组装机（BAM）复合物有效折叠OMP中的作用。我们发现，未能及时整合到膜中的β-桶底物通过Skp从BAM复合物中去除，从而允许清除停滞的Bam-OMP复合物。在将OMP从组装机器中置换后，Skp随后充当牺牲衔接蛋白以直接促进周质蛋白酶DegP降解缺陷OMP底物。我们得出结论，Skp通过直接介导来自Bam复合物的组装受损OMP底物的置换和降解来确保有效的β-桶折叠。
The outer membrane (OM) of gram-negative bacteria acts as a robust permeability barrier to enable cell survival in a wide variety of harsh environments. Crucial to OM integrity are β-barrel outer membrane proteins (OMPs) that are assembled into the membrane by the broadly conserved β-barrel assembly machine (Bam) complex. Here, we identify specific roles for the periplasmic chaperone Skp in functioning as a sacrificial adaptor protein to remove stalled substrates from the Bam complex, imposing an active quality control mechanism that ensures efficient assembly of nascent OMPs into the OM. This work identifies the molecular mechanism of the Skp/DegP functional relationship and clarifies the long-standing paradox of how substrate release from the high-affinity, long-lived Skp–OMP complex is achieved in vivo. The biogenesis of integral β-barrel outer membrane proteins (OMPs) in gram-negative bacteria requires transport by molecular chaperones across the aqueous periplasmic space. Owing in part to the extensive functional redundancy within the periplasmic chaperone network, specific roles for molecular chaperones in OMP quality control and assembly have remained largely elusive. Here, by deliberately perturbing the OMP assembly process through use of multiple folding-defective substrates, we have identified a role for the periplasmic chaperone Skp in ensuring efficient folding of OMPs by the β-barrel assembly machine (Bam) complex. We find that β-barrel substrates that fail to integrate into the membrane in a timely manner are removed from the Bam complex by Skp, thereby allowing for clearance of stalled Bam–OMP complexes. Following the displacement of OMPs from the assembly machinery, Skp subsequently serves as a sacrificial adaptor protein to directly facilitate the degradation of defective OMP substrates by the periplasmic protease DegP. We conclude that Skp acts to ensure efficient β-barrel folding by directly mediating the displacement and degradation of assembly-compromised OMP substrates from the Bam complex.
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