光谱法研究黄曲霉毒素B_1与人血清白蛋白的结合反应  

Fluorescence Spectroscopic Study of Interaction between Aflatoxin B1 and Human Serum Albumin

作　　者：江涛[1] 马良[1] 张宇昊[1] 王佳曼[1] 

机构地区：[1]西南大学食品科学学院,重庆400715

出　　处：《分析化学》2016年第1期54-60,共7页Chinese Journal of Analytical Chemistry

基　　金：国家重点基础研究发展计划(973计划)项目(No.2013CB127803);国家自然科学基金项目(No.31301476);中央高校基本科研业务费专项资金项目(No.2362014xk11)~~

摘　　要：黄曲霉毒素B1（AFB1）是目前发现的致癌能力最强的真菌毒素,严重危害人畜健康。人血清白蛋白（Human serum albumin,HSA）在结合、运输内源性和外源性等小分子物质方面具有重要的生理功能。研究AFB1与HSA的相互作用机理和作用过程,在分子毒理学上具有重要意义。本研究模拟在人体血液pH条件（pH 7.4,离子强度0.1 mol/L）,通过荧光淬灭、3D荧光法和圆二色谱（Circular dichroism,CD）等光谱方法研究AFB1与人血清蛋白的相互作用。结果表明,AFB1与HSA的内源荧光淬灭属于静态淬灭,AFB1-HSA在298,303,308和313 K4个温度条件下,结合常数均为10-4数量级,结合位点都约为1。根据Van＇t Hoff方程AFB1-HSA体系是熵增焓减的自发过程,分子间主要作用力为疏水作用和氢键。基于Forster＇s能量转移,得知AFB1与HSA结合距离为3.31 nm。竞争结合实验表明,AFB1结合在HSA的siteI位点上,靠近色氨酸Trp-214。通过3D荧光分析,AFB1的结合作用导致了HSA氨基酸残基微环境和二级构象发生变化。圆二色谱的分析结果表明,二者的结合使得HSA的α-螺旋含量增加。Aflatoxin B1（AFB1） is the most powerful cancer-causing mycotoxins,which is serious harmful to human and animal health.Human serum albumin has important physiological functions in the binding or transporting endogenous and exogenous ligands aspects.It＇s great significance in molecular toxicology of researching AFB1 and human serum albumin interaction mechanism.The interaction between AFB1 and human serum albumin（HSA） was investigated by fluorescence spectroscopy,circular dichroism and 3D fluorescence spectroscopy under the simulative physiological conditions（pH = 7.4,Ionic strength 0.1 mol/L）.Results showed that the main quenching mechanism between AFB1 and HSA was a static quenching process.At four different temperatures（298,303,308 and 313 K）,all the magnitude binding constants（K） were 104 and the number of binding sites（n） in the binary system was approximate to 1.According to Van＇t Hoff equation,the negative enthalpy change（△H-θ） and postive entropy change（△S-θ） values indicated that hydrophobic interaction and hydrogen bonding were the mainly interaction and force in the binding process.The binding distance（r） between the AFB1 and HSA was calculated to be 3.31 nm based on the theory of Forster＇s nonradiation energy transfer.The site marker displacement experiments suggested the location of AFB1 binding to HSA was site I,closely Trp-214.The 3D florescence revealed that the microenvironment of amino acid residues and the conformation of HSA were changed during the binding reaction.CD spectra revealed that the conformations of HSA were changed during the binding reaction with increasing in α-helix.

关 键 词：黄曲霉毒素B1 人血清白蛋白 结合反应 光谱法 

分 类 号：O657.3[理学—分析化学] TS201.6[理学—化学]