β/γ-Crystallins, the main structural protein in human lenses, have highly stable structure for keeping the lens transparent. Their mutations have been linked to congenital cataracts. In this study, we identified 10 new mutations of β/γ-crystallins in lens proteomic dataset of cataract patients using bioinformatic tools. Of these, two double mutants, S175G/H181Q of βB2-crystallin and P24S/S31G of γD-crystallin, were found mutations occurred in the largest loop linking the distant β-sheets in the Greek key motif. We selected these double mutants for identifying the properties of these mutations, employing biochemical assay, the identification of protein modifications with nanoUPLC-ESI-TOF tandem MS and examining their structural dynamics with hydrogen/deuterium exchange-mass spectrometry (HDX-MS). We found that both double mutations decrease protein stability and induce the aggregation of β/γ-crystallin, possibly causing cataracts. This finding suggests that both the double mutants can serve as biomarkers of congenital cataracts.

中文翻译：

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与白内障相关的βB2-晶体的新突变体S175G / H181Q和γD-晶体的P24S / S31G通过结构变化参与蛋白质聚集

β/γ-晶状体蛋白是人晶状体中的主要结构蛋白，具有高度稳定的结构，可保持晶状体透明。它们的突变与先天性白内障有关。在这项研究中，我们使用生物信息学工具在白内障患者的晶状体蛋白质组学数据集中确定了10个新的β/γ-晶状体蛋白突变。其中，发现两个双重突变体，βB2-crystallin的S175G / H181Q和γD-crystallin的P24S / S31G，在希腊钥匙基序中连接远处β-折叠的最大环中发生了突变。我们选择了这些双重突变体来鉴定这些突变体的特性，采用生化分析，使用nanoUPLC-ESI-TOF串联MS鉴定蛋白质修饰，并使用氢/氘交换质谱法（HDX-MS）检查其结构动力学。我们发现两个双重突变均会降低蛋白质稳定性并诱导β/γ-晶状体蛋白的聚集，可能引起白内障。这一发现表明两个双突变体都可以作为先天性白内障的生物标志物。