Department of Pharmacy, College of Pharmacy and Graduate School of Pharmaceutical Sciences, Ewha Womans University, Seoul 03760, Korea.
Department of Computer Science, Hanyang University, Seoul 04763, Korea.
Int J Mol Sci. 2020 Sep 5;21(18):6504. doi: 10.3390/ijms21186504.
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 cataracts. In this study, we identified 10 new mutations of β/γ-crystallins in lens proteomic dataset of cataract patients using bioinformatics tools. Of these, two double mutants, S175G/H181Q of βΒ2-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 cataracts.
β/γ-晶体蛋白是人类晶状体中的主要结构蛋白，其结构高度稳定，可保持晶状体透明。它们的突变与白内障有关。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鉴定了 10 种新的β/γ-晶体蛋白突变。其中，βΒ2-晶体蛋白的 S175G/H181Q 和 γD-晶体蛋白的 P24S/S31G 这两个双突变体，其突变发生在连接希腊钥匙模体中远端β-片层的最大环上。我们选择了这两个双突变体，通过生化分析、纳米 UPLC-ESI-TOF 串联 MS 鉴定蛋白质修饰以及氢/氘交换质谱（HDX-MS）检测其结构动力学，来确定这些突变的特性。我们发现这两种双突变都降低了蛋白质稳定性并诱导β/γ-晶体蛋白聚集，可能导致白内障。这一发现表明，这两种双突变体均可作为白内障的生物标志物。
