牦牛乳硬质干酪苦味肽RK7的抗氧化活性及其机制  

Antioxidant Activity and Mechanism of Yak Milk Hard Cheese Bitter Peptide RK7

作　　者：杨保军 梁琪[1,2] 宋雪梅 Yang Baojun;Liang Qi;Song Xuemei(College of Food Science and Engineering,Gansu Agricultural University,Lanzhou 730070;Functional Dairy Product Engineering Laboratory of Gansu,Lanzhou 730070)

机构地区：[1]甘肃农业大学食品科学与工程学院,兰州730070 [2]甘肃省功能乳品工程实验室,兰州730070

出　　处：《中国食品学报》2022年第8期40-50,共11页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家自然科学基金项目(31260383,31660468)。

摘　　要：目的:研究从牦牛乳硬质干酪中鉴定出苦味肽RK7的抗氧化活性及其机制。方法:运用生物信息学方法计算和分析RK7的理化和生物学特性,采用固相合成技术体外人工合成RK7并测定其抗氧化活性。借助分子对接工具,基于长寿酶家族sirtuins(SIRT1-7)和单磷酸腺苷激活的蛋白激酶(AMPK)介导的信号通路,研究RK7的抗氧化机制。结果:RK7的不稳定性指数为19.84;疏水性为42.86%;生物活性评分为0.404;小肠吸收能力(HIA)为0.9072+;血脑屏障穿透能力(BBB)为0.9701-;急性口腔毒性为0.6259;合成纯度为99.579%;当肽质量浓度在0.2~1.0 mg/mL范围内变化时,DPPH自由基清除率为12.26%~53.78%,ABTS自由基清除率为80.86%~87.86%;RK7与NAD+降解酶(3DZK)和胞质接头蛋白(2FLU)分别形成6个和5个氢键,3DZK的氨基酸残基Arg140、Asp147和2FLU的氨基酸残基Ser390、Asp394是其与RK7结合的重要活性位点。RK7和GSSH与3DZK和2FLU的结合表现出相似的分子机制,分别结合于3DZK的Site 1区域(X:-11.93,Y:-1.30,Z:0.38,R:10A)和2FLU的Site 2区域(X:8.33,Y:13.99,Z:20.41,R:9A)。研究结果为从分子水平分析、解释RK7的抗氧化性提供科学依据。Objective:The purpose of this experiment was to study the antioxidant activity and mechanism of the bitter peptide RK7 identified in yak milk hard cheese.Method:The bioinformatics methods were used to calculate and analyze the physicochemical properties and biological properties of RK7.The solid-phase synthesis technology was used to synthesize RK7 in vitro and detect its antioxidant activity.The molecular docking tool was used to study the antioxidant mechanism of RK7 based on the signal pathway mediated by longevity enzyme family sirtuins(SIRT1-7)and adenosine monophosphate-activated protein kinase(AMPK).The results showed that the instability index of RK7 was 19.84,the hydrophobicity was 42.86%,the biological activity score was 0.404,human intestinal aabsorption abilty(HIA)was 0.9072+,blood-brain barrier penetration abilty(BBB)was 0.9701-,acute oral toxicity was 0.6259,and the synthetic purity was 99.579%.When the peptide mass concentration changes in the range of 0.2-1.0 mg/mL,the scavenging rate of DPPH free radical was 12.26%-53.78%,and the scavenging rate of ABTS free radical was 80.86%-87.86%.RK7 formed 6 and 5 hydrogen bonds with NAD+degrading enzymes(3DZK)and cytoplasmic linker protein(2FLU)respectively.The amino acid residues Arg140 and Asp147 of 3DZK and the amino acid residues Ser390 and Asp394 of 2FLU were important active sites for binding to RK7.The combination of RK7 and GSSH with 3DZK and 2FLU showed similar molecular mechanisms,binding to the Site 1 area of 3DZK(X:-11.93,Y:-1.30,Z:0.38,R:10A)and the Site 2 area of 2FLU(X:8.33,Y:13.99,Z:20.41,R:9A),respectively.The above research provided a scientific basis for analyzing and explaining the antioxidant activity of RK7 at the molecular level.

关 键 词：牦牛乳硬质干酪 苦味肽 抗氧化活性 分子对接 

分 类 号：TS252.53[轻工技术与工程—农产品加工及贮藏工程]