大豆内源蛋白酶水解油体蛋白的机理及其对油脂体加工特性的影响

Soybean oil body is natural nanoscale raw material, which has great potential for application. In oil body extraction process, extrinsic protein in protein storage vacuole are adsorbed on the oil body surface, in which P34 is specific endogenous protease to hydrolyze 24 kDa and 18kDa oleosin and affect the processing properties of oil body emulsion. so it is urgent to investigate the mechanism of hydrolysis of soybean oleosin by P34. At present, domestic and foreign scholars have only studied the hydrolysis process of 24 kDa oleosin. However, the mechanism of hydrolysis of 18 kDa soybean oleosin by P34 has not been well revealed. In this study, soybean oil body concentrate is object of study, and the mechanism of hydrolysis of 18 kDa soybean oleosin by P34 will be researched. The hydrolysis product of 18 kDa oleosin is separated By RP-HPLC.The N-terminus sequence and molecular weight are identified by electrophoresis and HPLC to determine the hydrolysis site of 18 kDa oleosin.we will get the thermal dynamic data by using microcalorimetry of international advanced technology, to characterize scientificly and quantitatively the law of reaction conditions and changes.This will clarify the mechanism of hydrolysis of P34 18 kDa oleosin and reveal the influence on processing properties of oil body concentrate by hydrolysis. This research will perfect the mechanism of P34 hydrolysis of soybean oleosin, and provide theoretical support for domestic and foreign scholars to carry out microcalorimetric study of protease hydrolysis of soybean protein, and provide theoretical basis for the application of soybean oil body concentrate in food.

大豆油脂体为天然的纳米级原料，具有巨大应用潜力。油脂体提取过程，蛋白质贮存液泡中的蛋白质吸附在油脂体表面，其中特异性内源蛋白酶P34会水解24kDa和18kDa油体蛋白，影响油脂体乳液加工特性，因此P34水解大豆油体蛋白的机理急需解决。目前，国内外学者只研究了P34对24kDa油体蛋白的水解。P34水解18kDa油体蛋白的机理没有很好的揭示。本课题以大豆油脂体富集物为研究对象，研究P34水解18kDa油体蛋白，分离出18kDa的水解产物，通过测定水解产物N端序列和分子量来确定18kDa的水解位点；利用国际先进的微量热技术获得热动力学动态数据，科学、定量表征反应过程及变化规律，阐明P34水解18kDa的机理；揭示水解对油脂体加工特性的影响规律。该研究将完善P34水解大豆油体蛋白的作用机理，为开展蛋白酶水解大豆蛋白的微量热研究提供理论支持，为大豆油脂体富集物在食品中的应用提供理论基础。

大豆油脂体提取过程中蛋白质贮存液泡中的蛋白质会吸附在油脂体表面，其中特异性内源蛋白酶P34等会水解18kDa等油脂体蛋白，影响油脂体乳液加工特性。本课题采用微量热技术以及SDS-PAGE，Tricine-SDS-PAGE，MALDI-TOF/TOF，XRD，FT-IR，DSC,GC-MS等现代仪器分析方法，结合有机化学、物理化学等多学科理论研究了大豆内源蛋白酶水解大豆油脂体蛋白的机理及其对油脂体加工特性的影响规律。获得了不同酸碱度、温度、金属离子等条件下内源蛋白酶P34水解18kDa的热动力学参数等分子相互作用的完整热动力学信息，科学、定量表征了大豆内源蛋白酶水解油脂体蛋白的反应过程及规律；揭示了内源性蛋白酶作用下大豆油脂体蛋白水解对油脂体乳液粒径、ζ-电位、流变学性质和氧化稳定性影响的内在规律，并建立了有效控制模型；阐明了内源蛋白酶水解油脂体蛋白的机理以及内源蛋白酶水解油脂体蛋白对油脂体加工特性及产品质量的影响机制。为开展蛋白酶水解大豆蛋白的微量热研究提供了技术指导与理论支持，为大豆油脂体在食品中的应用及有效提升和控制大豆油脂体基食品品质提供了技术指导与理论基础。已在国内外重要期刊上发表论文11篇（其中SCI收录论文7篇），申请发明专利2项，授权专利1项，培养研究生7名。