The nascent polypeptide-associated complex modulates interactions between the signal recognition particle and the ribosome
The nascent polypeptide-associated complex modulates interactions between the signal recognition particle and the ribosome
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DOI：
10.1016/s0960-9822(02)00484-0
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发表时间：
1996-03-01
期刊：
影响因子：
9.2
通讯作者：
Walter, P
中科院分区：
文献类型：
--
作者：
Powers, T;Walter, P
背景：分泌蛋白共翻译靶向内质网膜的第一步涉及信号识别颗粒 (SRP) 的 54 kDa 亚基对从核糖体中出现的信号序列进行识别。最近有人提出，新生多肽相关复合物（NAG）通过调节核糖体新生链复合物、SRP 和内质网膜之间发生的相互作用来提高靶向保真度。目前尚不清楚 NAC 如何影响 SRP 功能。结果：我们使用免疫印迹实验来监测在 NAG 不存在和存在的情况下 SRP 和核糖体新生链复合物之间的相互作用。在不存在 NAG 的情况下，SRP 仅以高 sari 抗性方式与含有信号序列的核糖体结合，证实了 SRP 对信号序列的特异性，在较低盐浓度下观察到 SRP 与无信号核糖体新生链的结合；然而，与该复合物结合的 SRP 量与与缺乏新生链的核糖体结合的 SRP 量无法区分。因此，这种盐敏感结合可能是 SRP 和核糖体之间独立于新生链发生的相互作用的结果。由 SRP54 和 SRP RNA 组成的最小颗粒足以赋予与包含信号序列的核糖体的耐盐结合，而与缺乏新生链的核糖体的盐敏感结合需要所有 SRP 亚基。添加纯化的 NAG 可以抑制 SRP 的这种盐敏感结合。结论：根据我们的结果，我们定义了 SRP 和核糖体新生链复合物之间的两种不同的相互作用模式：SRP54 和信号序列之间的耐盐相互作用，以及 SRP 的其他成分和核糖体之间的盐敏感相互作用。我们得出的结论是，NAC 不会直接影响 SRP 的信号序列识别，而是负向调节 SRP 和核糖体本身之间发生的相互作用。这些结果是根据 SRP 和 NAC 在蛋白质靶向过程中调节彼此活性的模型进行讨论的。
Background: The first step in the co-translational targeting of secretory proteins to the endoplasmic reticulum membrane involves the recognition of signal sequences by the 54 kDa subunit of the signal recognition particle (SRP) as they emerge from the ribosome. It has recently been proposed that the nascent polypeptide-associated complex (NAG) contributes to the fidelity of targeting by modulating interactions that occur between the ribosome-nascent chain complex, the SRP and the endoplasmic reticulum membrane. Precisely how NAC influences SRP function is presently unclear.Results: We have used immunoblotting experiments to monitor interactions between the SRP and the ribosome-nascent chain complex, in the absence and presence of NAG. In the absence of NAG, SRP binds in a high-sari-resistant manner only to ribosomes that contain a signal sequence, confirming the specificity of SRP for signal sequences, Binding of SRP to signalless ribosome nascent chains is observed at lower salt concentrations; however, the amount of SRP bound to this complex is indistinguishable from that bound to ribosomes lacking nascent chains. Thus, this salt-sensitive binding is likely to be the result of interactions between SRP and the ribosome that occur independently of the nascent chain. A minimal particle consisting of SRP54 and SRP RNA is sufficient to confer salt-resistant binding to ribosomes that contain signal sequences, whereas all of the SRP subunits are required for salt-sensitive binding to ribosomes that lack nascent chains. This salt-sensitive binding by SRP is inhibited by the addition of purified NAG.Conclusions: Based on our results, we define two distinct modes of interaction between SRP and the ribosome-nascent chain complex: salt-resistant interactions between SRP54 and signal sequences, and salt-sensitive interactions between additional components of SRP and the ribosome. We conclude that NAC does not directly influence signal sequence recognition by SRP but, rather, that it negatively modulates interactions that occur between SRP and the ribosome itself, These results are discussed in terms of a model wherein SRP and NAC regulate each others' activity during protein targeting.