白内障相关人γD晶状体蛋白的热诱导变性  

Heat-induced denaturation of cataract-related humanγD-crystallin

作　　者：周鑫 李珍艳 李淑媛 张文博 王晨轩[1] ZHOU Xin;LI Zhenyan;LI Shuyuan;ZHANG Wenbo;WANG Chenxuan(Department of Biophysics and Structural Biology,State Key Laboratory of Common Mechanism Research for Major Diseases,Institute of Basic Medical Sciences CAMS,School of Basic Medicine PUMC,Beijing 100005,China)

机构地区：[1]中国医学科学院基础医学研究所,北京协和医学院基础学院,生物物理及结构生物学系,重大疾病共性机制研究全国重点实验室,北京100005

出　　处：《基础医学与临床》2025年第1期1-6,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(32201142)。

摘　　要：目的揭示人γD晶状体蛋白(HGD)与先天性白内障相关突变体蛋白(HGD P23T)的热诱导变性效应,对比野生型与突变体蛋白质在加热过程中结构变化的差异性。方法体外表达纯化HGD与HGD P23T,测定蛋白质内源性荧光强度和静态光散射光强随温度的变化,揭示HGD与HGD P23T依赖于温度的折叠与聚集结构变化规律。结果温度低于70℃时,HGD与HGD P23T的内源荧光质心波长随温度升高向长波长移动,荧光强度降低,蛋白质构象去折叠。HGD P23T比HGD的构象稳定性差。温度高于70℃时,静态光散射强度随温度显著上升,蛋白质受热聚集。相对于野生型,HGD P23T表现出更强的聚集趋势。结论加热破坏γD晶状体蛋白的折叠构象,诱导去折叠态蛋白质发生聚集。疾病相关P23T突变显著降低了γD晶状体蛋白的构象稳定性。Objective To reveal the thermally induced denaturation of wild-type humanγD-crystallin(HGD)and congenital cataract-related mutant(HGD P23T),and compare the differences in the structural changes between wild-type and mutants during a heating process.Methods HGD and HGD P23T were expressed and purified.The temperature-dependent intrinsic fluorescence intensity and static light scattering intensity of the protein samples were measured to reveal the temperature-dependent folding and aggregation structural changes of HGD and HGD P23T.Results When the temperature was below 70℃,the barycentric mean of the intrinsic fluorescence of HGD and HGD P23T shifted towards a longer wavelength with increasing temperature and the fluorescence intensity decreased indicating the unfolded protein conformations.The conformational stability of HGD P23T was weaker than that of HGD.When temperature was higher than 70℃,the static light scattering intensity increased significantly with temperature,indicating protein aggregation upon heating.Relative to the wild-type,HGD P23T showed a stronger aggregation potency.Conclusions Heating disrupts the folding conformation ofγD-crystallin,induces the unfolded protein to aggregate.The disease-associated P23T mutation significantly reduces the conformational stability ofγD-crystallin.

关 键 词：晶状体蛋白质 热稳定性 聚集 内源性荧光 静态光散射 

分 类 号：Q67[生物学—生物物理学]