Niwa Tatsuya, Nakazawa Koki, Hoshi Kensuke, Tadakuma Hisashi, Ito Koichi, Taguchi Hideki
School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan.
Cell Biology Center, Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan.
Front Mol Biosci. 2022 Aug 25;9:891128. doi: 10.3389/fmolb.2022.891128. eCollection 2022.
Co-translational protein folding is one of the central topics in molecular biology. In , trigger factor (TF) is a primary chaperone that facilitates co-translational folding by directly interacting with nascent polypeptide chains on translating ribosomes. In this study, we applied fluorescence correlation spectroscopy (FCS), which can analyze the diffusion properties of fluorescent molecules by measuring the fluctuations of the fluorescent intensity, to investigate the interaction between TF and a nascent chain on translating ribosomes both and . The FCS analysis with a reconstituted cell-free translation system revealed that the interaction of fluorescently labeled TF with a nascent chain depended on the emergence of the nascent chain from the ribosome exit tunnel, and this interaction was not inhibited by excess amounts of other chaperones. Furthermore, the translation-dependent interaction between GFP-fused TFs and nascent chains was also observed in living cells. The FCS-based approach established here could be an effective method to investigate the dynamics of other ribosome-associated chaperones besides TF.
共翻译蛋白质折叠是分子生物学的核心主题之一。在[具体内容缺失]中，触发因子（TF）是一种主要的伴侣蛋白，它通过与正在翻译的核糖体上的新生多肽链直接相互作用来促进共翻译折叠。在本研究中，我们应用荧光相关光谱法（FCS），该方法可通过测量荧光强度的波动来分析荧光分子的扩散特性，以研究TF与正在翻译的核糖体上的新生链之间在[具体内容缺失]和[具体内容缺失]时的相互作用。对重组无细胞翻译系统进行的FCS分析表明，荧光标记的TF与新生链的相互作用取决于新生链从核糖体出口通道的出现，并且这种相互作用不受过量其他伴侣蛋白的抑制。此外，在活[具体内容缺失]细胞中也观察到了绿色荧光蛋白融合的TF与新生链之间依赖于翻译的相互作用。这里建立的基于FCS的方法可能是研究除TF之外的其他核糖体相关伴侣蛋白动力学的有效方法。
