Mechanistic details of the signal recognition particle (SRP)-mediated insertion of membrane proteins have been described from decades of in vitro biochemical studies. However, the dynamics of the pathway inside the living cell remain obscure. By combining in vivo single-molecule tracking with numerical modeling and simulated microscopy, we have constructed a quantitative reaction–diffusion model of the SRP cycle. Our results suggest that the SRP–ribosome complex finds its target, the membrane-bound translocon, through a combination of three-dimensional (3D) and 2D diffusional search, together taking on average 750 ms. During this time, the nascent peptide is expected to be elongated only 12 or 13 amino acids, which explains why, in Escherichia coli , no translation arrest is needed to prevent incorrect folding of the polypeptide in the cytosol. We also found that a remarkably high proportion (75%) of SRP bindings to ribosomes occur in the cytosol, suggesting that the majority of target ribosomes bind SRP before reaching the membrane. In combination with the average SRP cycling time, 2.2 s, this result further shows that the SRP pathway is capable of targeting all substrate ribosomes to translocons.

中文翻译：

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活大肠杆菌细胞中 SRP 通路的时空动力学

数十年的体外生化研究描述了信号识别颗粒 (SRP) 介导的膜蛋白插入的机制细节。然而，活细胞内通路的动力学仍然不清楚。通过将体内单分子跟踪与数值建模和模拟显微镜相结合，我们构建了 SRP 循环的定量反应-扩散模型。我们的结果表明，SRP-核糖体复合物通过三维 (3D) 和 2D 扩散搜索的组合找到其目标，即膜结合转位子，平均耗时 750 毫秒。在此期间，新生肽预计只会延长 12 或 13 个氨基酸，这就解释了为什么，在大肠杆菌, 不需要翻译停滞来防止胞质溶胶中多肽的不正确折叠。我们还发现 SRP 与核糖体结合的比例非常高 (75%) 发生在胞质溶胶中，这表明大多数目标核糖体在到达膜之前结合 SRP。结合平均 SRP 循环时间 2.2 秒，该结果进一步表明 SRP 途径能够将所有底物核糖体靶向转位子。