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Thermal-induced Unfolding of β-Crystallin and Disassembly of its Oligomers Revealed by Temperature-Jump Time-Resolved Infrared Spectroscopy
Shan-shan Li,Ying-ying Yu,De-yong Li,Xiao-chuan He,Yong-zhen Bao*,Yu-xiang Weng*
Author NameAffiliationE-mail
Shan-shan Li Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China  
Ying-ying Yu Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases, Key Laboratory of Vision Loss and Restoration, Ministry of Education, Department of Ophthalmology, Peking University People's Hospital, Beijing 100044, China  
De-yong Li Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China  
Xiao-chuan He Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China  
Yong-zhen Bao* Beijing Key Laboratory of Diagnosis and Therapy of Retinal and Choroid Diseases, Key Laboratory of Vision Loss and Restoration, Ministry of Education, Department of Ophthalmology, Peking University People's Hospital, Beijing 100044, China drbaoyz@sina.com 
Yu-xiang Weng* Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China yxweng@aphy.iphy.ac.cn 
Abstract:
β-Crystallins are the major structural proteins existing in the vertebrate lens, and their conformational stability is critical in maintaining the life-long transparency and refraction index of the lens. Seven subunits of β-crystallins naturally assemble into various heteroge-neous oligomers with different sizes. Here, we systematically investigated the thermal sta-bility of the different secondary structures present in β-crystallins and then the dynamic process for the thermal-induced unfolding of β-crystallins by Fourier transform infrared spectroscopy-monitored thermal titration and temperature-jump nanosecond time-resolved IR difference absorbance spectra. Our results show that the N-terminal anti-parallel β-sheets in β-crystallin are the most unstable with a transition midpoint temperature at 36.0±2.1 oC, leading to the formation of an intermediate consisting vastly of random coil structures. This intermediate structure is temporally assigned to that of the monomer generated by the thermal-induced disassembly of β-crystallin oligomers with a transition midpoint tempera-ture of 40.4±0.7 oC. The global unfolding of β-crystallins that leads to denaturation and aggregation indicated by the formation of intermolecular anti-parallel β-sheets has a transi-tion midpoint temperature determined as 72.4±0.2 oC. Temperature-jump time-resolved IR absorbance difference spectroscopy analysis further reveals that thermal-induced unfolding of β-crystallins occurs firstly in the anti-parallel β-sheets in the N-terminal domains with a time constant of 50 ns
Key words:  β-Crystallin, Protein dynamical structure, Temperature-jump, Time-resolved IR spectrum
FundProject:
热诱导β晶状体蛋白去折叠及寡聚体解聚过程的脉冲升温时间分辨红外光谱
李姗姗,余盈盈,李得勇,何小川,鲍永珍*,翁羽翔*
摘要:
通过变温傅里叶变换红外光谱和脉冲升温时间分辨红外光谱,系统地研究了β晶状体蛋白不同二级结构的热稳定性,以及热诱导β晶状体蛋白去折叠的动力学过程.结果表明,β晶状体蛋白N端的β反平行折叠热稳定性最低(转变的中点温度为36.0±2.1 oC),该结构的破坏导致了一种富含无规卷曲结构的β晶状体蛋白中间体形成, 认为该中间体可能由β晶状体寡聚体解聚后形成的单体,单体形成的中点温度为40.4±7 πC.β晶状体蛋白全局去折叠引发蛋白变性聚集的转变的中点温度为72.4±0.2 oC.脉冲升温时间分辨红外光谱进一步揭示出β晶状体蛋白的热诱导去折叠开始于N端的β反平行折叠结构,该结构去折叠所需时间约为50 ns
关键词:  β晶状体蛋白,蛋白质动态结构,脉冲升温,时间分辨红外光谱
DOI:10.1063/1674-0068/26/06/739-746
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