PDF(Pubmed)
Abstract：
Eye lens α- and β-/γ-crystallin proteins are not replaced after fiber cell denucleation and maintain lens transparency and refractive properties. The exceptionally high (∼400-500 mg/mL) concentration of crystallins in mature lens tissue and multiple other factors impede precise characterization of β-crystallin interactions, oligomer composition, size, and topology. Native ion mobility-mass spectrometry is used here to probe β-crystallin association and provide insight into homo- and heterooligomerization kinetics for these proteins. These experiments include separation and characterization of higher-order β-crystallin oligomers and illustrate the unique advantages of native IM-MS. Recombinantly expressed βB1, βB2, and βA3 isoforms are found to have different homodimerization propensities, and only βA3 forms larger homooligomers. Heterodimerization of βB2 with βA3 occurs ∼3 times as fast as that of βB1 with βA3, and βB1 and βB2 heterodimerize less readily. Ion mobility experiments, molecular dynamics simulations, and PISA analysis together reveal that observed oligomers are consistent with predominantly compact, ring-like topologies.
摘要：
眼晶状体α-和β-/γ-晶状体蛋白在纤维细胞去核后不会被取代，并保持晶状体的透明度和屈光特性。在成熟的晶状体组织中晶体蛋白的异常高（〜400-500mg/mL）浓度和多种其他因素阻碍了β-晶体蛋白相互作用的精确表征，低聚物成分,尺寸,和拓扑。这里使用天然离子迁移-质谱来探测β-晶状体蛋白缔合，并提供对这些蛋白质的均聚和杂低聚动力学的了解。这些实验包括高阶β-晶状体蛋白低聚物的分离和表征，并说明了天然IM-MS的独特优势。发现重组表达的βB1，βB2和βA3亚型具有不同的同二聚化倾向，并且只有βA3形成更大的均低聚物。βB2与βA3的异源二聚化的发生速度是βB1与βA3的3倍，而βB1和βB2的异源二聚化较不容易。离子迁移实验,分子动力学模拟，和PISA分析一起显示，观察到的寡聚体与主要是紧密的，环状拓扑。
