Chen Xiuqi, Hilser Vincent J, Kaiser Christian M
CMDB Graduate Program, Johns Hopkins University, Baltimore, MD, USA.
Department of Biology, Johns Hopkins University, Baltimore, MD, USA.
Nat Commun. 2025 Jul 24;16(1):6833. doi: 10.1038/s41467-025-61398-6.
Cytosolic proteins begin to fold co-translationally as soon as they emerge from the ribosome during translation. These early co-translational steps are crucial for overall folding and are guided by an intricate network of interactions with molecular chaperones. Because cellular co-translational folding is challenging to detect, its timing and progression remain largely elusive. To quantitatively define co-translational folding in live cells, we developed a high-throughput method that we term "Arrest Peptide Profiling" (AP Profiling). Combining AP Profiling with single-molecule experiments, we delineate co-translational folding for a set of GTPase domains with similar structures, defining how topology shapes folding pathways. Genetic ablation of nascent chain-binding chaperones results in discrete and localized folding changes, highlighting how functional redundancy among chaperones is achieved by distinct engagement with the nascent protein. Our work provides a window into cellular folding pathways of structurally intricate proteins and paves the way for systematic studies of nascent protein folding at exceptional resolution and throughput.
在翻译过程中，胞质蛋白一旦从核糖体中出现，就开始进行共翻译折叠。这些早期的共翻译步骤对于整体折叠至关重要，并由与分子伴侣的复杂相互作用网络引导。由于细胞共翻译折叠难以检测，其时间和进程在很大程度上仍然难以捉摸。为了定量定义活细胞中的共翻译折叠，我们开发了一种高通量方法，我们称之为“捕获肽谱分析”（AP谱分析）。将AP谱分析与单分子实验相结合，我们描绘了一组结构相似的GTPase结构域的共翻译折叠，确定了拓扑结构如何塑造折叠途径。新生链结合伴侣的基因消融导致离散和局部的折叠变化，突出了伴侣之间的功能冗余是如何通过与新生蛋白质的不同结合来实现的。我们的工作为深入了解结构复杂蛋白质的细胞折叠途径提供了一个窗口，并为以超高分辨率和通量对新生蛋白质折叠进行系统研究铺平了道路。
