青刺果ACE抑制肽的分离纯化、结构鉴定及其体外活性评价
Isolation,Identification,Structure Characterization and in Vitro Activity Evaluation of ACE Inhibitory Peptides from Prinsepia utilis
张璐 1甘雨嫣 1钟玉旺 1黎依艳 1范江平 2王雪峰2
作者信息
- 1. 云南农业大学食品科学技术学院 昆明 650201
- 2. 云南农业大学食品科学技术学院 昆明 650201;云南省畜产品加工工程技术研究中心 昆明 650201
- 折叠
摘要
以具有血管紧张素转化酶(ACE)抑制活性的青刺果蛋白酶解物为研究对象,采用超滤、强阴离子交换层析分离纯化ACE抑制肽,液相色谱-串联质谱鉴定其肽序列.采用傅里叶红外光谱、酶反应抑制剂动力学、MTT试验和分子对接技术解析其二级结构特征、体外活性以及与ACE的结合机制.结果表明,分子质量＜3 ku的超滤组分活性较好(IC50=0.380mg/mL),离子交换层析后以F-a组分活性最好(IC50=0.159mg/mL).质谱鉴定出4条肽序列,通过生物信息学分析确定肽PGDVF为潜在的ACE抑制肽[IC50=(0.56±0.1)mmol/L].二级结构分析表明PGDVF由α-螺旋(20.28％)、β-折叠(6.21％)、β-转角(31.55％)和无规则卷曲(41.85％)构成,其抑制模型为非竞争性抑制,肽质量浓度小于1 mg/mL时,对HepG2细胞无毒性.分子对接显示PGDVF可通过氢键、疏水作用与ACE紧密结合,从而有效抑制ACE活性.研究结果可为青刺果降压肽的开发利用提供参考.
Abstract
In this study,ACE inhibitory peptides were isolated and purified by ultrafiltration and strong anion exchange chromatography,and their peptide sequences were identified by LC-MS/MS.Fourier transform infrared spectroscopy,en-zyme reaction inhibitor kinetics,MTT assay and molecular docking technology were used to analyze the secondary struc-ture characteristics,in vitro activity and binding mechanism with ACE.The results showed that the activity of＜3 ku ultrafiltration fraction was better(IC50 value:0.380 mg/mL),and the activity of F-a fraction was the best(IC50 value:0.159 mg/mL)after ion exchange chromatography.Four peptide sequences were identified by LC-MS/MS,and bioinformatics analysis further confirmed that the peptide PGDVF was a potential ACE inhibitory peptide with an IC50 val-ue of(0.56±0.1)mmol/L.the secondary structure analysis showed that PGDVF was composed of α-helix,β-sheet,β-turn and random coil,and its inhibition model may be mixed.The peptide concentration less than 1 mg/mL was basical-ly non-toxic to HepG-2 cells.Molecular docking showed that PGDVF could be closely combined with ACE enzyme through hydrogen bond and hydrophobic force,thus effectively inhibiting ACE activity.In summary,this study can pro-vide reference for the development and utilization of Prinsepia utilis decompression peptide.
关键词
青刺果/ACE抑制肽/液相色谱-串联质谱/二级结构/分子对接Key words
Prinsepia utilis/angiotensin-I converting enzyme inhibitory peptide/liquid chromatography-tandem mass spectrometry/secondary structure/molecular docking引用本文复制引用
基金项目
国家自然科学基金地区项目(31960462)
云南省基础研究计划面上项目(2019FB052)
云南省农业联合专项重点项目(202101BD070001-013)
云南省青年科技人才托举工程项目()
出版年
2024
NETL
NSTL
万方数据