Trigger factor forms a protective shield for nascent polypeptides at the ribosome
Trigger factor forms a protective shield for nascent polypeptides at the ribosome
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DOI：
10.1074/jbc.m512345200
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发表时间：
2006-03-10
影响因子：
4.8
通讯作者：
Bukau, B
中科院分区：
文献类型：
--
作者：
Hoffmann, A;Merz, F;Bukau, B
在原核生物中，核糖体相关触发因子是新合成的多肽从核糖体出口通道出来时遇到的第一个伴侣。然而，触发因子对新生多肽的影响尚不清楚。在这里，我们分析了触发因子在核糖体上屏蔽新生多肽的潜力。在大肠杆菌为基础的体外转录/翻译系统中合成了一组不同来源、大小和折叠状态的新生多肽，并测试了对非特异性蛋白酶k降解的敏感性。在缺乏触发因子的情况下，暴露在核糖体出口通道外的新生多肽迅速降解，除非它们折叠成致密结构域。触发因子的存在，以及缺乏肽基脯氨酸异构酶结构域的触发因子片段，抵消了所有未折叠的新生多肽的降解。这种保护功能是核糖体拴系的触发因子所特有的，因为无论是非核糖体触发因子还是在体内折叠过程中与触发因子合作的DnaK系统，都没有显示出类似的保护效率。此外，触发因子的屏蔽并不局限于新生链的短片段，对于暴露在核糖体外高达41 kDa的大型非天然新生多肽也很明显。我们认为触发因子通过屏蔽核糖体上的新生多肽来支持生产的新生折叠，从而防止过早的降解或聚集过程。触发因子提供的这种受保护的环境可能对大型多结构域蛋白有效地折叠成其天然状态特别重要。
In prokaryotes, the ribosome-associated Trigger Factor is the first chaperone newly synthesized polypeptides encounter when they emerge from the ribosomal exit tunnel. The effects that Trigger Factor exerts on nascent polypeptides, however, remain unclear. Here we analyzed the potential of the Trigger Factor to shield nascent polypeptides at the ribosome. A set of arrested nascent polypeptides differing in origin, size, and folding status were synthesized in an Escherichia coli-based in vitro transcription/translation system and tested for sensitivity to degradation by the unspecific protease proteinase K. In the absence of Trigger Factor, nascent polypeptides exposed outside the ribosomal exit tunnel were rapidly degraded unless they were folded into a compact domain. The presence of Trigger Factor, as well as a Trigger Factor fragment lacking its peptidyl-prolyl isomerase domain, counteracted degradation of all unfolded nascent polypeptides tested. This protective function was specific for ribosome-tethered Trigger Factor, since neither non-ribosomal Trigger Factor nor the DnaK system, which cooperates with Trigger Factor in the folding process in vivo, revealed a comparable efficiency in protection. Furthermore, shielding by Trigger Factor was not restricted to short stretches of nascent chains but was evident for large, non-native nascent polypeptides exposing up to 41 kDa outside the ribosome. We suggest that Trigger Factor supports productive de novo folding by shielding nascent polypeptides on the ribosome thereby preventing untimely degradation or aggregation processes. This protected environment provided by Trigger Factor might be particularly important for large multidomain proteins to fold productively into their native states.