带电多糖对油体蛋白有机溶剂诱导下疏水聚集的“保护-去保护”机制研究

油体蛋白为植物种子储油亚细胞结构—油脂体的界面稳定蛋白，极具开发成为高效表面活性剂的潜质。而在提取过程中有机溶剂沉淀引发其强烈疏水聚集，低溶解度成为应用开发与基础研究的限制性因素。油体蛋白增溶问题核心在于减少疏水聚集以保持其天然的两亲构象。本课题拟筛选具有显著维持油体蛋白界面构象与活性的带电多糖，构建油体蛋白与多糖静电相互作用模型，依次建立“多糖结构”-“界面分子相互作用”-“界面结构”-“油体蛋白聚集与构象转变”之间的明确构效关系，为制备基于油体蛋白的新型乳化剂提供直接理论支撑。综合物理化学表征、界面特性、蛋白质表达技术与蛋白质组学等系列方法，阐述多糖结构与静电复合参数在油茶种子油体蛋白“去折叠”与“再折叠”过程中的分子调控机制，构建一种基于食品组分之间相互作用而减少蛋白质在有机溶剂变性中的疏水聚集，促进再次溶水恢复原构象，实现简单、安全、温和的蛋白质构象“保护-去保护”策略。

Oil body associated proteins (OAPs) are the interfacial structural proteins of the oil body, which is the subcellular structure of plant seeds for oil storage. They have great potentials to be developed as an efficient emulsifier or foam stabilizer. However, during the extraction process, the hydrophobic aggregation of OAPs induced by organic solvents resulting in precipitation. The very low solubility limits their structural characterization, functional properties, and applications in the food industry. The key problem of increasing the solubility and keeping their functional properties is to maintain the original amphiphilic conformation of OAPs. In this project, we will first select the charged polysaccharides which could maintain the interfacial conformation of OAPs, construct the model of electrostatic interaction between charged polysaccharides and OAPs, and state a clear structure-activity relationship among the structure of polysaccharides, electrostatic binding, interfacial structure, and the aggregation behavior of OAPs. The results would provide theoretical parameters for developing new emulsifiers based on OAPs. Through combining a series of experiments including characterization of physical and chemical properties, observation of interfacial structure, biosynthesis of oleosin by protein expression, and identification of proteomics, the mechanism of polysaccharides on the modification of camellia OAPs structure and their electrostatic binding status during the unfolding and refolding process of the oleosin at the oil-water interface is illustrated. Besides, we hope to establish a method based on the interaction between food components with proteins for reducing the hydrophobic aggregation induced by organic solvents. It is a kind of protection and deprotection strategy, which promotes the recovery of the oleosin conformation in the aqueous phase with simple, safe and temperate ways.