绿豆胰脂肪酶抑制肽的制备、鉴定及抑制机制

Preparation, identification and inhibitory mechanism of pancreatic lipase inhibitory peptide from mung bean protein

中文关键词:  绿豆蛋白  蛋白酶解  胰脂肪酶抑制  脂肪代谢  分子对接

英文关键词:Mung bean protein  Proteolysis  Pancreatic lipase inhibition  Lipid metabolism, Molecular docking

中文摘要:

      以绿豆蛋白为原料,利用超声辅助蛋白酶酶解法制备胰脂肪酶抑制肽(Mung bean protein pancreatic lipase inhibitory peptide,MPLIPs),并对其抑制胰脂肪酶活性的机制进行了研究。探究了蛋白酶的种类、酶解时间、温度、蛋白酶用量等因素对水解度和胰脂肪酶抑制率的影响,胃蛋白酶的酶解产物对胰脂肪酶活性抑制率最高；利用响应面法对工艺条件进行了优化,结果如下：酶解温度为40℃,酶解时间3 h,蛋白酶添加量为13000 U/g,所得到MPLIPs对胰脂肪酶抑制率为68.34%,水解度为13.02%；MPLIPs浓度为600 μg/mL时其半抑制浓度IC50为442.18 μg/mL；对绿豆蛋白酶解液进行脱色、脱盐,超滤处理,结果表明分子量在450～3000 Da的MPLIPs的胰脂肪酶抑制活性最高；利用LC-MS/MS鉴定出35条绿豆活性肽段,通过Peptide Ranker网站进行虚拟筛选,得出5条具有活性潜力的肽段；利用分子对接技术研究了GIQRGAAGAELLRSWSFGM肽对胰脂肪酶的抑制机制,该肽与胰脂肪酶(1ETH)结合能较低,结合稳定性好,其作用力主要为氢键和疏水作用。这表明MPLIPs对胰脂肪酶具有良好的抑制作用,可用于调节脂肪代谢类功能性食品研究与开发。

英文摘要:

      Mung bean protein pancreatic lipase inhibitory peptide (MPLIPs) was prepared by ultrasonic- assisted enzymatic hydrolysis using mung bean protein as the raw material and the mechanism inhibiting pancreatic lipase activity was studied. The effects of experimental factors such as protease types, hydrolysis time, temperature and the amount of protease on hydrolysis degree and the inhibition rate of pancreatic lipase were investigated, the enzymatic hydrolysate of pepsin exhibited the highest inhibitory rate on pancreatic lipase activity. The optimal conditions were obtained by response surface methodology (RSM) , the results as follows: the enzymatic hydrolysis temperature as 40℃, the enzymatic hydrolysis time as 3 h, the amount of protease was 13000 U/g. The inhibitory rate of pancreatic lipase was 68.34%, the degree of hydrolysis was 13.02% at these conditions, and the IC50 of MPLIPs was 442.18 μg /mL at the concentration of 600 μg / mL . After decolorization, desalination, and ultrafiltration of mung bean protein hydrolysate, MPLIPs with molecular weights between 450–3000 Da exhibited higher pancreatic lipase inhibitory activity. 35 active peptides were identified by LC-MS/MS, and 5 active peptides were screened by Peptide Ranker. The inhibition mechanism of GIQRGAAGAELLRSWSFGM peptide on pancreatic lipase was studied by molecular docking. The binding energy of GIQRGAAGAELLRSWSFGM peptide to pancreatic lipase was lower, their binding is relatively stable, with the main forces being hydrogen bonds and hydrophobic interactions. This indicates that MPLIPs have a good inhibitory effect on pancreatic lipase and can be used in the development of functional foods for regulating lipid metabolism.