带电短肽T_(7)+、T_(6)-对α-淀粉酶活性和构象的影响  

Effect of Charged Short Peptides T_(7)+and T_(6)-on the Activity and Structure of α-Amylase

作　　者：杨晓钰 李传博 刘春莹 孙付保[4] 窦少华[1,2,3] Yang Xiaoyu;Li Chuanbo;Liu Chunying;Sun Fubao;Dou Shaohua(School of Life and Health,Dalian University,Dalian 116622,Liaoning;Liaoning Marine Microbial Engineering and Technology Research Centre,Dalian 116622,Liaoning;Key Laboratory of Animal Immunity,Dalian 116622,Liaoning;School of Bioengineering,Jiangnan University,Wuxi 214122,Jiangsu)

机构地区：[1]大连大学生命健康学院,辽宁大连116622 [2]辽宁省海洋微生物工程技术研究中心,辽宁大连116622 [3]大连市动物免疫重点实验室,辽宁大连116622 [4]江南大学生物工程学院,江苏无锡214122

出　　处：《中国食品学报》2024年第7期14-23,共10页Journal of Chinese Institute Of Food Science and Technology

基　　金：辽宁省教育厅高校基本科研面上项目(JYTMS20230376);辽宁省自然科学基金项目(2014020134)。

摘　　要：利用带不同电荷的短肽T_(7)+、T_(6)-,探讨正电荷短肽T_(7)+和负电荷短肽T_(6)-对α-淀粉酶的活性和构象的影响。将短肽加入α-淀粉酶体系后,测定其相对酶活、米氏常数(Km)、活化能、Zeta电位等指标。为了更清晰地观察不同带电量的短肽对α-淀粉酶构象的影响,通过紫外吸收光谱、荧光光谱、同步荧光光谱、圆二色光谱进一步考察α-淀粉酶构象的变化。结果表明,将带电短肽加入α-淀粉酶的酶促反应中,正电荷短肽T7+抑制α-淀粉酶酶活,当短肽质量浓度为10^(-7)g/mL时,α-淀粉酶相对酶活降低2.40%,同时也使α-淀粉酶与可溶性淀粉的亲和力降低,活化能升高,Zeta电位增加。负电荷短肽T_(6)-促进α-淀粉酶酶活,当短肽质量浓度为10^(-7)g/mL时,α-淀粉酶相对酶活升高1.40%,同时也使α-淀粉酶与可溶性淀粉的亲和力升高,活化能降低,Zeta电位增加。光谱分析表明,不同带电短肽使α-淀粉酶的紫外-可见吸收光谱、荧光光谱以及二级结构发生不同程度的改变。这表明不同带电短肽可以发挥与电场相似的作用,即电场在作用于蛋白质时,产生的电场力促使蛋白质构象发生改变,人工设计合成短肽所带的表面电荷影响酶表面电荷的分布,进而导致酶的活性和构象发生改变。In this experiment,differently charged short peptides T_(7)+and T_(6)-were used to investigate the mechanism of the change in α-amylase activity and conformation by the positively charged short peptide T_(7)+and the negatively charged short peptide T_(6)-.The changes of α-amylase activity and conformation were verified by the determination of relative enzyme activity,Mie's constant(Km),activation energy and zeta potential after the addition of short peptides to α-amylase system.In order to observe more clearly the effects of different charged short peptides on the conformation of α-amylase,UV absorption spectra,fluorescence spectra,simultaneous fluorescence spectra and circular dichroism spectra were also introduced to further investigate the conformational changes of α-amylase.The results showed that the addition of the charged short peptide into the enzymatic reaction of α-amylase,the positively charged short peptide T7+inhibited theα-amylase enzyme activity,and the relative enzyme activity of α-amylase decreased by 2.40% when the short peptide concentration was 10^(-7) g/mL,and also decreased the affinity of α-amylase with soluble starch,increased the activation energy and increased the zeta potential.The negatively charged short peptide T_(6)-promoted theα-amylase enzyme activity,and when the short peptide concentration was 10^(-7) g/mL,the relative enzyme activity ofα-amylase increased by 1.40%,and also made the affinity ofα-amylase with soluble starch increase,the activation energy decrease and the zeta potential increase.Spectroscopic analysis showed that different charged short peptides changed the UV-visible absorption spectrum,fluorescence spectrum and secondary structure of α-amylase to different degrees.It was demonstrated that different charged short peptides could play a role similar to that of electric field,the electric field force generated by the electric field when acting on the proteins contributed to the conformational change of the proteins,and the surface charge of the artifici

关 键 词：Α-淀粉酶 带电短肽 相对酶活 构象 

分 类 号：TS201.25[轻工技术与工程—食品科学]