Improved enzymatic accessibility of peanut protein isolate pre-treated using thermosonication
Improved enzymatic accessibility of peanut protein isolate pre-treated using thermosonication
复制标题
使用热超声预处理提高花生分离蛋白的酶促可达性
DOI：
10.1016/j.foodhyd.2019.02.050
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发表时间：
2019-08-01
影响因子：
10.7
通讯作者：
Akhtar, Mahmood
中科院分区：
文献类型：
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作者：
Chen, Lin;Ettelaie, Rammile;Akhtar, Mahmood
热超声预处理被用于增强胰蛋白酶诱导的花生分离蛋白（PPI）的水解作用。采用响应面法优化热超声条件（包括功率输出和温度），在475.0瓦、72℃时获得了最高水解度（7.16%）。SDS - PAGE分析表明，在该优化条件下，热超声处理的PPI（TS - PPI）中主要组成蛋白花生球蛋白的酶可及性与未处理的PPI或超声处理的PPI（475瓦，30℃；S - PPI）相比显著提高，导致水解产物的蛋白质溶解度显著增加。利用差示扫描量热法、内源荧光发射光谱法、傅里叶变换红外光谱法和硫黄素 - T（ThT）荧光测定法研究了未处理的PPI、S - PPI和TS - PPI的蛋白质变性和构象特征。研究发现，热量对超声使花生蛋白变性具有显著的叠加效应，导致蛋白质构象发生显著变化。TS - PPI的特征是出现高比例的平行分子间β - 折叠，并且在与ThT结合时荧光显著增强，这表明热超声诱导的蛋白质去折叠和聚集可能导致TS - PPI中形成纤维状蛋白质聚集体，而非S - PPI中形成的球状蛋白质聚集体。因此，TS - PPI的蛋白质构象似乎比未处理的PPI或S - PPI更伸展、更灵活，因而更容易被蛋白酶作用。这项研究表明，热超声预处理可能是一种高效且可行的技术，可提高球状蛋白质的酶可及性，为蛋白质水解产物带来显著的功能优势。
Thermosonication pre-treatment was used to enhance the pancreatin-induced proteolysis of peanut protein isolate (PPI). Response surface methodology was applied to optimize the thermosonication conditions (including power-output and temperature), and the highest degree of hydrolysis (7.16%) was obtained at 475.0 W, 72 degrees C. SDS-PAGE analysis showed that at this optimized condition, the enzymatic accessibility of the major constitutive protein arachin in thermosonicated PPI (TS-PPI) was substantially improved compared to that in untreated PPI or sonicated PPI (475 W, 30 degrees C; S-PPI), resulting in a remarkable increase in protein solubility for the hydrolysates. Protein denaturation and conformation profiles of untreated PPI, S-PPI and TS-PPI were investigated using differential scanning calorimetry, intrinsic fluorescence emission spectroscopy, Fourier transform infra-red spectroscopy and thioflavin-T (ThT) fluorescence assay. It was found that heat could present a markedly additive effect to ultrasound on denaturing peanut proteins, leading to significant changes in protein conformation. TSPPI was characterized by the appearance of high proportion of parallel intermolecular beta-sheets and a strong fluorescence enhancement upon binding to ThT, suggesting that the protein unfolding and aggregation induced by thermosonication probably resulted in the formation of fibril protein aggregates in TS-PPI rather than spherical protein aggregates formed in S-PPI. As a result, the protein conformation of TS-PPI appeared to be more unfolded and flexible than that of untreated PPI or S-PPI, and therefore was more easily accessible to protease. This study shows that thermosonication pre-treatment could be a highly effective and feasible technique to improve the enzymatic accessibility of globular proteins, producing prominent functional benefits for the protein hydrolysates.