【作者】 贺舒雯； 韩鹏薇； 杜雨凌； 杨赛杰； 李虹辉； 吴昊； 文李； 程云辉；

【Author】 He Shuwen;Han Pengwei;Du Yuling;Yang Saijie;Li Honghui;Wu Hao;Wen Li;Cheng Yunhui;Department of Food and Biological Engineering,Changsha University of Science & Technology;

【机构】 长沙理工大学食品与生物工程学院；

【摘要】 食源性抗氧化肽作为一种天然抗氧化剂,具有安全、活性好、易吸收等优点,对人体健康具有重要作用。Keap1/Nrf2信号通路是机体重要的抗氧化应激通路,在维持机体内氧化还原动态平衡过程中发挥关键作用。本研究以克氏原螯虾虾壳为原料,利用胰蛋白酶、胃蛋白酶、碱性蛋白酶、风味蛋白酶、木瓜蛋白酶等5种蛋白酶分别进行酶解,以ABTS、DPPH、超氧阴离子和羟自由基清除率为指标,筛选出抗氧化活性较高的蛋白酶水解产物;通过LC-MS/MS鉴定虾壳肽序列,结合肽-Keap1分子对接模拟筛选潜在抗氧化肽并利用SPR分析其结合亲和力。结果显示:碱性蛋白酶水解产物具有较高的氨基酸含量,小于1 ku的分子量较多,且ABTS、DPPH和超氧阴离子自由基清除能力较强;碱性蛋白酶水解产物中鉴定得到1600条肽序,通过在线网站预测和分子对接发现Keap1蛋白与筛选的30条潜在抗氧化肽均能结合,经SPR检测发现WL5和WI5肽与Keap1蛋白具有较强的结合亲和力。本研究初步揭示了虾壳肽WL5和WI5可以通过与Keap1蛋白结合抑制Keap1/Nrf2相互作用发挥抗氧化作用,为抗氧化肽的研究和应用奠定基础,同时也对克氏原螯虾副产物的高值化利用提供参考。更多还原

【Abstract】 As a natural antioxidant,food-derived antioxidant peptide has the advantages of safety,activity,and easy absorption,which plays an important role in human health.keap1/Nrf2 signaling pathway is an important antioxidant stress pathway in the body and plays a key role in maintaining the dynamic balance of redox in the body.In this study,five proteases,including trypsin,pepsin,alkaline protease,flavor protease,and papain,were used as raw materials for enzymatic digestion of Procambarus clarkii shells,and the protease hydrolysis products with high antioxidant activity were screened by using ABTS,DPPH,superoxide anion and hydroxyl radical scavenging rate as indicators.The peptide sequence of Procambarusclarkii shells was identified by LC-MS/MS,combined with peptide-Keapl molecular docking simulation to screen potential antioxidant peptides and analyzed their binding affinity by SPR.The results showed that the alkaline protease hydrolysis products had high amino acid content,more with a molecular weight of less than 1 ku,and strong scavenging ability of ABTS,DPPH and superoxide anion radicals;1600 peptide sequences were identified in the alkaline protease hydrolysis products,and the Keapl protein was found to bind to 30 potential antioxidant peptides by online website prediction and molecular docking.The peptides WL5 and WI5 were found to have a strong binding affinity to Keapl protein by SPR.This study initially revealed that the shrimp shell peptides WL5 and WI5 could inhibit the Keap1/Nrf2 interaction by binding to Keapl protein,laying a foundation for the research and application of antioxidant peptides,and also providing a reference for the high-value utilization of crayfish by-products.更多还原