生姜蛋白酶水解产物中二肽基肽酶-Ⅳ抑制肽的虚拟筛选及活性分析  

Virtual Screening and Activity Evaluation of Dipeptidyl Peptidase-ⅣInhibitory Peptides from Ginger Protease Hydrolysates

作　　者：刘芬娣 李婧铭 陈洪娇 李雪岩 刘洋[2] 刘伟 LIU Fendi;LI Jingming;CHEN Hongjiao;LI Xueyan;LIU Yang;LIU Wei(Beijing Language University Hospital,Beijing 100083,China;Department of Chemistry of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 102488,China;Department of Traditional Chinese Medicine and Rehabilitation,Beijing Health Vocational College,Beijing 101101,China)

机构地区：[1]北京语言大学医院,北京100083 [2]北京中医药大学中药学院,北京102488 [3]北京卫生职业学院中药与康复系,北京101101

出　　处：《现代食品科技》2024年第5期34-42,共9页Modern Food Science and Technology

基　　金：国家自然科学基金面上项目(8197131426)。

摘　　要：根据生姜蛋白酶的水解特异性,从其水解物中虚拟筛选出高活性肽段,进行体外活性验证和体内吸收入血成分研究。利用Peptide Ranker和分子对接虚拟筛选出高活性肽段,建立体外酶促反应体系测定活性肽对二肽基肽酶-Ⅳ(DPP-Ⅳ)的抑制活性(IC50),并且在此基础上,引入表面等离子体共振(SPR)技术深入研究DPP-Ⅳ与活性肽的相互作用,并开展大鼠灌胃实验研究活性肽在胃肠道消化系统中的稳定性。通过计算机模拟,虚拟筛选得到Gly-Pro-Ser-Gly-Pro-Hyp-Gly-Pro-Hyp-Gly-Pro-Hyp-Gly(GPSGPXGPXGPXG)DPP-Ⅳ抑制肽。体外酶促实验表明GPSGPXGPXGPXG对DPP-Ⅳ具有较强的抑制作用,IC50为457.3μmol/L,SPR实验进一步表明二者之间具有快速结合和快速解离的动力学过程,但是大鼠体内实验提示,该多肽无法维持胃肠道稳定性,不是以原型形式被消化道吸收,而是部分以二肽和三肽的形式进入血液循环系统,包括Pro-Hyp、Gly-Pro-Hyp和Pro-Hyp-Gly等寡肽。该研究表明筛选得到的活性肽口服经消化道吸收后无法保持结构完整性,会被体内的肽酶水解,这为后续进行体内的药代动力学和药效学研究提供理论支撑。Based on the hydrolysis specificity of ginger protease,highly bioactive peptide segments were virtually screened from the hydrolysates,followed by validation of in vitro activity and investigation of in vivo absorption into blood.First,highly active peptide segments were virtually screened through Peptide Ranker and molecular docking.Next,an in vitro enzyme-catalyzed reaction system was established to determine the inhibitory activity(IC50)of active peptides against dipeptidyl peptidase-Ⅳ(DPP-Ⅳ).Based on this,surface plasmon resonance(SPR)was introduced to investigate the interactions between DPP-Ⅳand active peptides.Finally,oral administration experiments in rats were conducted to study the stability of active peptides in the gastrointestinal tract.Through computer simulation,the DPP-Ⅳinhibitory peptide Gly-Pro-Ser-Gly-Pro-Hyp-Gly-Pro-Hyp-Gly-Pro-Hyp-Gly(GPSGPXGPXGPXG)was obtained via virtual screening.Moreover,the in vitro enzyme-catalyzed experiments showed that GPSGPXGPXGPXG has strong inhibitory effects against DPP-Ⅳ,with an IC50 of 457.3µmol/L.SPR experiments further revealed fast binding and dissociation kinetics between the two peptides.However,in vivo experiments in rats suggested that GPSGPXGPXGPXG is not stable in the gastrointestinal tract and it is not absorbed intact,but it is partially absorbed into the circulatory system in the form of dipeptides and tripeptides,including oligopeptides like Pro-Hyp,Gly-Pro-Hyp,and Pro-Hyp-Gly.This study demonstrates that the bioactive peptides obtained via screening cannot maintain their structural integrity after oral absorption through the digestive tract and are hydrolyzed by peptidases in the body.The findings provide theoretical support for subsequent pharmacokinetic and pharmacodynamic studies in vivo.

关 键 词：生姜蛋白酶 虚拟筛选 药靶结合动力学 表面等离子体共振技术 

分 类 号：TS201.2[轻工技术与工程—食品科学]