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学科主题临床医学
Oriented immobilisation of histidine-tagged protein and its application in exploring interactions between ligands and proteins
Zhao, Xinfeng1,4; Li, Qian1; Xiao, Chaoni1; Zhang, Yajun1; Bian, Liujiao1; Zheng, Jianbin2; Zheng, Xiaohui1; Li, Zijian3; Zhang, Youyi3; Fan, Taiping4
关键词Protein-druginteractions Affinity Chromatography Oriented Immobilisation Histidine Tag Beta(2)-adrenoceptor
刊名ANALYTICAL AND BIOANALYTICAL CHEMISTRY
2014-05-01
DOI10.1007/s00216-014-7723-x
406期:12页:2975-2985
收录类别SCI
文章类型Article
WOS标题词Science & Technology
类目[WOS]Biochemical Research Methods ; Chemistry, Analytical
研究领域[WOS]Biochemistry & Molecular Biology ; Chemistry
关键词[WOS]HUMAN SERUM-ALBUMIN ; AFFINITY-CHROMATOGRAPHY ; LIQUID-CHROMATOGRAPHY ; BINDING ; BIOSENSOR ; BETA(2)-ADRENOCEPTOR ; ANTIBODIES ; RECEPTOR ; SURFACE ; DRUGS
英文摘要

A method based on reaction with a diazonium salt was developed to immobilise oriented His-tagged protein onto silica gel. The binding efficiency of the phenylamine-group-coated gel was determined to be 65 %, providing a binding capacity of His-tagged protein up to the gram level. Using His-tagged beta(2)-adrenoceptor (beta(2)-AR) as a probe, we developed a new mathematical model to elucidate the interactions between the receptor and five ligands (methoxyphenamine, terbutaline, salbutamol, tulobuterol and fenoterol). These drugs proved to only have one type of binding site on the immobilised beta(2)-AR, yielding higher association constants and numbers of binding sites than random attachment assays. The association constants determined by the new model positively correlated to the values from a radioligand binding method, with a regression equation of y = 1.75x -aEuro parts per thousand 7.18 and a correlation coefficient of 0.9807. The oriented method resulted in a high binding capacity and quantitative immobilisation of the His-tagged protein. The proposed model can be used to determine the interactions between the ligands and the immobilised protein with the advantages of drug and time saving.

语种英语
WOS记录号WOS:000334431300024
项目编号21005060 ; IRT1174 ; 2013KCT-24
资助机构National Natural Sciences Foundation of China ; Program for Changjiang Scholars and Innovative Research Team in University ; project for Innovative Research Team of Research and Technology of Shaanxi Province
引用统计
被引频次:8[WOS]   [WOS记录]     [WOS相关记录]
文献类型期刊论文
条目标识符http://ir.bjmu.edu.cn/handle/400002259/56049
专题北京大学第三临床医学院
作者单位1.Univ Cambridge, Dept Pharmacol, Cambridge CB2 1PD, England
2.NW Univ Xian, Coll Life Sci, Key Lab Resource Biol & Biotechnol Western China, Minist Educ, Xian 710069, Peoples R China
3.NW Univ Xian, Inst Analyt Sci, Xian 710069, Peoples R China
4.Peking Univ, Hosp 3, Inst Vasc Med,Minist Hlth, Key Lab Cardiovasc Mol Biol & Regulatory Peptide, Beijing 100083, Peoples R China
推荐引用方式
GB/T 7714
Zhao, Xinfeng,Li, Qian,Xiao, Chaoni,et al. Oriented immobilisation of histidine-tagged protein and its application in exploring interactions between ligands and proteins[J]. ANALYTICAL AND BIOANALYTICAL CHEMISTRY,2014,406(12):2975-2985.
APA Zhao, Xinfeng.,Li, Qian.,Xiao, Chaoni.,Zhang, Yajun.,Bian, Liujiao.,...&Fan, Taiping.(2014).Oriented immobilisation of histidine-tagged protein and its application in exploring interactions between ligands and proteins.ANALYTICAL AND BIOANALYTICAL CHEMISTRY,406(12),2975-2985.
MLA Zhao, Xinfeng,et al."Oriented immobilisation of histidine-tagged protein and its application in exploring interactions between ligands and proteins".ANALYTICAL AND BIOANALYTICAL CHEMISTRY 406.12(2014):2975-2985.
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