核桃抗氧化肽的酶法制备、鉴定及其对脂氧合酶的抑制机理  

Enzymatic Preparation and Identification of Walnut Antioxidant Peptides and Their Inhibitory Mechanism on Lipoxygenase

作　　者：洪佳伟 肖柳柳 王爱琳 李闯 朱西平 顾千辉 谢婷婷 薛正莲[1] HONG Jiawei;XIAO Liuliu;WANG Ailin;LI Chuang;ZHU Xiping;GU Qianhui;XIE Tingting;XUE Zhenglian(School of Biological and Food Engineering,Anhui Polytechnic University,Wuhu 241000,China;Three Squirrels Co.,Ltd.,Wuhu 241000,China)

机构地区：[1]安徽工程大学生物与食品工程学院,安徽芜湖241000 [2]三只松鼠股份有限公司,安徽芜湖241000

出　　处：《食品与生物技术学报》2024年第6期112-118,共7页Journal of Food Science and Biotechnology

基　　金：安徽省重点研发计划项目(202104a06020003)。

摘　　要：作者以核桃粕为原料制备抗氧化肽,用胰酶酶解核桃蛋白质,以超氧阴离子自由基清除率为主要指标优化酶解条件,分离纯化酶解产物。通过LC-MS/MS技术鉴定多肽序列,通过最低对接结合能、蛋白质稳定指数等参数筛选抗氧化肽。结果表明,最优酶解条件下,酶解液的超氧阴离子自由基清除率为26.57%。酶解液经Smartdex G-50分离纯化得到4个组分,在相同质量浓度下,D-4组分超氧阴离子自由基清除率最高。对D-4组分进行序列鉴定,经计算机虚拟筛选推测R.AGIQFPVG.R为D-4组分抗氧化活性的主要贡献物。该肽段和脂氧合酶最低对接结合能为-21933.85 J/mol,蛋白质预测结果为稳定蛋白质。对接结果表明,R.AGIQFPVG.R和脂氧合酶非活性位点Glu186结合,说明该肽段主要起非竞争性抑制作用,该研究为天然抗氧化剂的筛选提供参考。To enhance the utilization of walnut meal,walnut protein was enzymatically hydrolyzed by trypsin and the purified hydrolysates were separated to analyze the antioxidant peptides.The preparation conditions were optimized by using superoxide anion free-radical scavenging as the main indicator.The purified components were sequenced by LC-MS/MS and the peptides were screened by docking binding energy and protein stability index.Under optimum hydrolysis conditions,the superoxide anion free-radical scavenging rate of hydrolysate was 26.57%.Altogether four hydroly-sate fractions were separated by Smartdex G-50,among which,the hydrolysate,namely D-4 displayed the highest scavenging rate at the same concentration.The sequence of D-4 component was identified and R.AGIQFPVG.R was presumed to be the main bearer of antioxidant activity,as determined by virtual computer screening.Through molecular docking with lipoxygenase,it was found that R.AGIQFPVG.R was able to form hydrogen bonds with inactive center of lipoxygenase with a binding energy of-21933.85 J/mol.The docking results showed that R.AGIQFPVG.R bound to the lipoxygenase inactive site via Glu186,indicating that the peptide acted mainly as a non-competitive inhibitor.This study conducted an enzymatic hydrolysis of walnut meal to prepare antioxidant peptides to provide a reference research for the screening of natural antioxidants.

关 键 词：核桃 抗氧化肽 分离纯化 分子对接 生物信息学 酶解 

分 类 号：TS201.1[轻工技术与工程—食品科学]