Data and scripts from: Temperature-dependent interactions explain normal and inverted solubility in a γD-crystallin mutant

Protein crystal production is a major bottleneck in the structural characterisation of proteins. To advance beyond large-scale screening, rational strategies for protein crystallization are crucial. Understanding how chemical anisotropy (or patchiness) of the protein surface due to the variety of amino acid side chains in contact with solvent, contributes to protein-protein contact formation in the crystal lattice is a major obstacle to predicting and optimising crystallization. The relative scarcity of sophisticated theoretical models that include sufficient detail to link collective behaviour, captured in protein phase diagrams, and molecular level details, determined from high-resolution structural information is a further barrier. Here we present two crystals structures for the P23T+R36S mutant of γD-crystallin, each with opposite solubility behaviour -- one melts when heated, the other when cooled. When combined with the protein phase diagram and a tailored patchy particle model we show that a single temperature dependent interaction is sufficient to stabilise the inverted solubility crystal. This contact, at the P23T substitution site, relates to a genetic cataract and reveals at a molecular level, the origin of the lowered and retrograde solubility of the protein. Our results show that the approach employed here may present an alternative strategy for the rationalization of protein crystallization.

蛋白质晶体制备是蛋白质结构表征的核心瓶颈。为突破大规模筛选的局限，合理的蛋白质结晶策略至关重要。理解由与溶剂接触的各类氨基酸侧链所引发的蛋白质表面化学各向异性（或称斑块化），如何促进晶格内蛋白质-蛋白质接触的形成，是预测并优化结晶过程的核心障碍。当前，尚缺乏能够充分关联两类关键信息的复杂理论模型：一类是蛋白质相图中所捕捉到的集体行为，另一类是通过高分辨率结构信息所确定的分子层面细节，这进一步成为研究的又一壁垒。本文报道了γD晶状体蛋白（γD-crystallin）P23T+R36S突变体的两种晶体结构，二者呈现截然相反的溶解行为：其一在升温时发生溶解，其二在降温时发生溶解。结合蛋白质相图与定制化斑块粒子模型，我们证实仅需单一的温度依赖性相互作用，即可稳定具有逆温溶解特性的晶体。该相互作用位点恰为P23T突变位点，与遗传性白内障密切相关，并从分子层面揭示了该蛋白质溶解度下降及逆行溶解特性的根源。本研究结果表明，本文所采用的研究思路可为蛋白质结晶的合理化设计提供一种全新的可行策略。