Liu Shao-Chuang, Ying Yi-Lun, Li Wei-Hua, Wan Yong-Jing, Long Yi-Tao
State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University Nanjing 210023 P. R. China
Department of Chemistry, East China University of Science and Technology Shanghai 200237 P. R. China.
Chem Sci. 2021 Jan 4;12(9):3282-3289. doi: 10.1039/d0sc06106a.
A fundamental question relating to protein folding/unfolding is the time evolution of the folding of a protein into its precisely defined native structure. The proper identification of transition conformations is essential for accurately describing the dynamic protein folding/unfolding pathways. Owing to the rapid transitions and sub-nm conformation differences involved, the acquisition of the transient conformations and dynamics of proteins is difficult due to limited instrumental resolution. Using the electrochemical confinement effect of a solid-state nanopore, we were able to snapshot the transient conformations and trace the multiple transition pathways of a single peptide inside a nanopore. By combining the results with a Markov chain model, this new single-molecule technique is applied to clarify the transition pathways of the β-hairpin peptide, which shows nonequilibrium fluctuations among several blockage current stages. This method enables the high-throughput investigation of transition pathways experimentally to access previously obscure peptide dynamics, which is significant for understanding the folding/unfolding mechanisms and misfolding of peptides or proteins.
一个与蛋白质折叠/去折叠相关的基本问题是蛋白质折叠成其精确界定的天然结构的时间演化过程。正确识别过渡构象对于准确描述动态蛋白质折叠/去折叠途径至关重要。由于涉及快速转变和亚纳米级别的构象差异，受限于仪器分辨率，获取蛋白质的瞬态构象和动力学较为困难。利用固态纳米孔的电化学限制效应，我们能够捕捉单个肽在纳米孔内的瞬态构象，并追踪其多条转变途径。通过将结果与马尔可夫链模型相结合，这种新的单分子技术被用于阐明β-发夹肽的转变途径，该肽在几个阻塞电流阶段之间表现出非平衡波动。这种方法能够通过实验高通量地研究转变途径，以了解之前模糊不清的肽动力学，这对于理解肽或蛋白质的折叠/去折叠机制以及错误折叠具有重要意义。
