退火促进变性蛋白的正确折叠
黄清艺#，杨留艳#，沈庆涛*
（1.南方科技大学生命科学学院，广东 深圳 518055；2.青岛海洋科技中心海洋生物学与生物技术功能实验室，山东 青岛 266237；3.南方科技大学生物电镜研究院，广东 深圳 518055；4. 山东大学微生物技术研究院，山东 青岛 266237；5.崂山国家实验室，山东 青岛 266237）
摘 要 退火技术是冶金工业常用的热处理工艺，通过对金属、合金和半导体材料进行先加热后降温的处理，可以同步化其微观结构，改善其物理和化学性质。前期，作者率先把退火引入生物大分子的结构生物学研究，实现对生物大分子构象的同步化，进而提高冷冻电镜解析分辨率。为了拓宽退火的应用，本文以绿色荧光蛋白（EGFP）为研究对象，对热变性的EGFP进行退火处理，发现退火处理的EGFP展现出更快的复性速度和更高的复性效率，这一发现有利于深入理解蛋白质折叠的机制，也为利用退火稳定和修复变性蛋白提供了理论基础和实验支撑。
关键词 退火；蛋白质复性；负染电镜；绿色荧光蛋白
中图分类号： Q518.4；Q336 文献标识码：ADoi:10.3969/j.issn.1000-6281.2025.01.006
Annealing promotes the refolding of denatured EGFPs
HUANG Qingyi#, YANG Liuyan#, SHEN Qingtao
（1. School of Life Sciences, Department of Chemical Biology, Southern University of Science and Technology, Shenzhen Guangdong 518055；2. Laboratory for Marine Biology and Biotechnology, Qingdao Marine Science and Technology Center, Qingdao Shandong 266237；3. Institute for Biological Electron Microscopy, Southern University of Science and Technology, Shenzhen Guangdong 518055；4. State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Shandong University, Qingdao Shandong 266237；5. Laoshan Laboratory, Qingdao Shandong 266237, China）
Abstract Annealing is widely utilized in metallurgy that synchronizes the microstructure of metals, alloys, and semiconductor materials to improve their physical and chemical properties. For the first time, we applied annealing onto biological macromolecules, which synchronizes them into a minimum-energy state after the tandem heating and cooling process. To expand the applications of annealing, we applied annealing on heat-denatured enhanced green fluorescent protein (EGFP) and found that annealed EGFP exhibited higher renaturation efficiency and faster renaturation speed. Our study helps deepen the understanding of protein folding mechanisms and provides a theoretical foundation and experimental support for stabilizing and repairing denatured proteins using annealing.
Keywords annealing；protein refolding；negative staining electron microscopy；green fluorescent protein
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