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首页> 外文期刊>The protein journal >Unfolding and Refolding of Bovine alpha-Crystallin in Urea and Its Chaperone Activity.
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Unfolding and Refolding of Bovine alpha-Crystallin in Urea and Its Chaperone Activity.

机译：尿素中牛α-晶体的折叠和重折叠及其伴侣活性。

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We undertook an unfolding and refolding study of alpha(L)-crystallin in presence of urea to explore the breakdown and formation of various levels of structure and to find out whether the breakdown of various levels of structure occurs simultaneously or in a hierarchal manner. We used various techniques such as circular dichroism, fluorescence spectroscopy, light scattering, polarization to determine the changes in secondary, tertiary, and quaternary structure. Unfolding and refolding occurred through a number of intermediates. The results showed that all levels of structure in alpha(L)-crystallin collapsed or reformed simultaneously. The intermediates that occurred in the 2-4 M urea concentration range during unfolding and refolding differed from each other in terms of the polarity of the tryptophan environment. The ANS binding experiments revealed that refolded alpha(L)-crystallin had higher number of hydrophobic pockets compared to native one. On the other hand, polarity of these pockets remained same as that of the native protein. Both light scattering and polarization measurements showed smaller oligomeric size of refolded alpha(L)-crystallin. Thus, although the secondary structural changes were almost reversible, the tertiary and quaternary structural changes were not. The refolded alpha(L)-crystallin had more exposed hydrophobic sites with increased binding affinity. The refolded form also showed higher chaperone activity than native one. Since the refolded form was smaller in oligomeric size, some buried hydrophobic sites were available. The higher chaperone activity of lower sized oligomer of alpha(L)-crystallin again revealed that chaperone activity was dependent on hydrophobicity and not on oligomeric size.
机译：我们进行了尿素存在下α（L）-晶状体蛋白的展开和再折叠研究，以探索各种结构水平的分解和形成，并找出各种结构水平的分解是同时发生还是以分层方式发生。我们使用了多种技术，例如圆二色性，荧光光谱，光散射，偏振来确定二级，三级和四级结构的变化。展开和重新折叠通过许多中间体发生。结果表明，α（L）-晶状体蛋白的所有结构水平同时崩溃或重新形成。在展开和再折叠过程中，在2-4 M尿素浓度范围内出现的中间体在色氨酸环境的极性方面彼此不同。 ANS结合实验表明，与天然的相比，重折叠的alpha（L）-晶状体蛋白具有更多的疏水口袋。另一方面，这些口袋的极性与天然蛋白质的极性保持相同。光散射和偏振测量均显示重折叠的alpha（L）-晶状蛋白的寡聚体尺寸较小。因此，尽管二级结构变化几乎是可逆的，但第三级和第四级结构变化却不可逆。重新折叠的α（L）-晶状蛋白具有更多的暴露疏水位点，结合亲和力增加。重新折叠的形式还显示出比天然形式更高的伴侣活性。由于重折叠形式的寡聚体尺寸较小，因此可以使用一些掩埋的疏水位点。较低尺寸的α（L）-晶状蛋白低聚物的较高分子伴侣活性再次表明，分子伴侣活性取决于疏水性，而不取决于寡聚体大小。

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