Oligomerization and phase transitions in aqueous solutions of native and truncated human βB1-crystalline
Oligomerization and phase transitions in aqueous solutions of native and truncated human βB1-crystalline
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DOI：
10.1021/bi048419f
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发表时间：
2005-02-01
期刊：
影响因子：
2.9
通讯作者：
Benedek, GB
中科院分区：
文献类型：
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作者：
Annunziata, O;Pande, A;Benedek, GB
人βB1-晶状体蛋白是一种主要的眼晶状体蛋白，随着年龄的增长，它在N端发生截短。通过研究天然BetaB1和截断BetaB1DeltaN41(模拟体内年龄相关的截断)，我们定量地确定了截断对BetaB1水溶液齐聚和相变性质的影响。齐聚研究表明，BetaB1DeltaN41蛋白之间的吸引能比BetaB1蛋白的吸引能大约10%。我们发现BetaB1DeltaN41水溶液经历了两种不同类型的相变。第一个相变涉及细棒状组件的初始形成，然后这些组件演化形成晶体。棒状聚集体形成的诱导时间对齐聚反应很敏感。第二个相变可以被描述为液-液相分离(LLP)，并伴随着富含蛋白质的相中的凝胶。我们将这一过程称为异相凝胶化。在BetaB1水溶液中没有观察到这两个相变。然而，在加入聚乙二醇(PEG)后，我们也观察到BetaB1的异相凝胶化。我们的聚乙二醇实验使我们能够估计BetaB1和BetaB1DeltaN41之间的相分离温度差异。这一差异与我们在齐聚研究中发现的吸引能增加是一致的。我们的工作表明，截断是一种导致白内障的修饰，因为它有利于蛋白质凝聚和随后形成的光散射元件，并强调了BetaB1的N末端在维持晶状体透明度方面的重要性。
Human betaB1-crystallin is a major eye-lens protein that undergoes in vivo truncation at the N-terminus with aging. By studying native betaB1 and truncated betaB1DeltaN41, which mimics an age-related in vivo truncation, we have determined quantitatively the effect of truncation on the oligomerization and phase transition properties of betaB1 aqueous solutions. The oligomerization studies show that the energy of attraction between the betaB1DeltaN41 proteins is about 10% greater than that of the betaB1 proteins. We have found that betaB1DeltaN41 aqueous solutions undergo two distinct types of phase transitions. The first phase transition involves an initial formation of thin rodlike assemblies, which then evolve to form crystals. The induction time for the formation of rodlike assemblies is sensitive to oligomerization. The second phase transition can be described as liquid-liquid phase separation (LLPS) accompanied by gelation within the protein-rich phase. We refer to this process as heterogeneous gelation. These two phase transitions are not observed in the case of betaB1 aqueous solutions. However, upon the addition of poly(ethylene glycol) (PEG), we observe heterogeneous gelation also for betaB1. Our PEG experiments allow us to estimate the difference in phase separation temperatures between betaB1 and betaB1DeltaN41. This difference is consistent with the increase in energy of attraction found in our oligomerization studies. Our work suggests that truncation is a cataractogenic modification since it favors protein condensation and the consequent formation of light scattering elements, and highlights the importance of the N-terminus of betaB1 in maintaining lens transparency.