目的:采用酶解法制备克氏原螯虾壳蛋白水解物(Procambarus clarkii shell protein hydrolysates,PCSPHs),并分析其体外降糖降脂活性及肽序.方法:分别采用胃蛋白酶、碱性蛋白酶、胰蛋白酶、风味蛋白酶和木瓜蛋白酶水解制备不同虾壳蛋白水解物,分析其体外降糖降脂活性和肽序列;运用 Peptide Ranker及BIOPEP-UWM网站在线分析,再以核受体PPARγ配体结合区域的晶体结构作为靶点,使用Autodock vina进行分子对接模拟,获得具有潜在降糖降脂活性的虾壳肽.结果:胃蛋白酶水解物(PEP-PCSPHs)对α-淀粉酶和α-葡萄糖苷酶活性具有较强的抑制作用,IC50值分别为(5.42±0.05),(7.11±1.01)mg/mL;胰蛋白酶水解物(TRY-PCSPHs)对胰脂肪酶活性具有最强的抑制能力,IC5.值为(4.71±1.12)mg/mL,且对甘氨胆酸钠表现出最好的体外结合效果.此外,经质谱鉴定PEP-PCSPHs和TRY-PCSPHs中分别得到3 391,2 086条肽序;通过在线网站预测和分子对接筛选出多条均能与PPARγ结合的降糖降脂虾壳活性肽(PCSAPs).结论:酶解克氏原螯虾壳制备的虾壳蛋白水解物具有潜在的降糖降脂活性,可能改善糖脂代谢紊乱.
Abstract
Objective:The PCSPHs were prepared by enzymatic hydrolysis of Procambarus clarkii shells,and their hypoglycemic and lipid-lowering activities in vitro were evaluated and peptide sequence were analyzed.Methods:Different crayfish shell proteolysates were prepared by hydrolysis of pepsin,alcalase protease,trypsin,flavor protease and papain,and their in vitro hypoglycemic and lipid-lowering activities were evaluated and peptide sequences were determined.The peptides sequence of Procambarus clarkii shells was identified by LC-MS/MS.Taking the crystal structure of the nuclear receptor PPARγ ligand binding region as the target,Autodock vina was used to simulate molecular docking to obtain crayfish shell peptides with potential hypoglycemic/lipid-lowering activities.Results:The PEP-PCSPHs had significant inhibitory effects on α-amylase andα-glucosidase activity,with IC50 values of(5.42±0.05)mg/mL and(7.11±1.01)mg/mL,respectively.The TRY-PCSPHs had the strongest inhibitory effect on pancreatic lipase activity,with an IC50 of(4.71±1.12)mg/mL,and exhibited the best in vitro binding effects on sodium glycinocholate.In addition,3 391 peptide sequences were identified in pepsin hydrolysates and 2 086 peptide sequences were identified in trypsin hydrolysates,and multiple hypoglycemic/lipid-lowering crayfish shell active peptides that could bind to PPARγ were screened through online website prediction and molecular docking.Conclusion:The shrimp shell peptides prepared by enzymatic hydrolysis of crayfish shells have potential hypoglycemic and lipid-lowering activities,which may play a role in improving glucose and lipid metabolism disorders.
关键词
克氏原螯虾壳/蛋白水解物/PPARγ/分子对接/降糖降脂活性
Key words
Procambarus clarkii shells/protein hydrolysate/PPARγ/molecular docking/hypoglycemic and lipid-lowering activity
维普
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