物理化学学报 >> 2004, Vol. 20 >> Issue (02): 216-220.doi: 10.3866/PKU.WHXB20040223

研究简报 上一篇    下一篇

糖蛋白-凝集素自组装构筑有序膜及在酶电极的应用

李扬眉;江秀明;陈志春;傅水玉;林贤福   

  1. 浙江大学化学系,杭州 310027;杭州职业技术学院化工系,杭州 310018;杭州师范学院化学系,杭州 310012
  • 收稿日期:2003-08-05 修回日期:2003-09-19 发布日期:2004-02-15
  • 通讯作者: 林贤福 E-mail:llc123@css.zju.edu.cn

Construction of Ordered Films of Horseradish Peroxidase through Lectin-glycogenReaction and Its Application in Enzyme Electrode

Li Yang-Mei;Jiang Xiu-Ming;Chen Zhi-Chun;Fu Shui-Yu;Lin Xian-Fu   

  1. Chemistry Department of Zhejiang University, Hangzhou 310027; Chemical Engineering Department of Hangzhou Professional Technical College, Hangzhou 310018; Chemistry Department of Hangzhou Teachers College, Hangzhou 310012
  • Received:2003-08-05 Revised:2003-09-19 Published:2004-02-15
  • Contact: Lin Xian-Fu E-mail:llc123@css.zju.edu.cn

摘要: 利用糖蛋白-凝集素的识别作用交替沉积伴刀豆球蛋白(Con A)与辣根过氧化物酶(HRP)制备酶自组装多层膜,用原子力显微镜(AFM)观测了自组装膜的表面形貌、表面粗糙度; AFM和椭圆偏振研究测定了自组装膜的厚度.结果表明, Con A和HRP膜厚分别为9.0和4.6 nm,与两者的晶体衍射结果一致,说明生物识别自组装方法能很好地保持分子的原有形态.以亚甲蓝(MB)溶液为介体,用循环伏安法测定了表面修饰了三层(Con A/HRP)自组装膜的金电极对H2O2的电化学催化还原作用,在H2O2浓度为0.2~1.0 mmol•L-1时,响应电流对H2O2浓度变化成线性,酶电极灵敏度为24.0 mA•mol-1•L,表观米氏常数为4.2 mmol•L-1.

关键词: 辣根过氧化物酶, 伴刀豆球蛋白, 表面形貌, 自组装膜, AFM, 酶电极

Abstract: The multilayer films of horseradish peroxidase (HRP) were fabricated by layer-by-layer assembly of HRP and concanavalin A (Con A) through lectin-glycogen reaction. The surface topography and roughness were characterized by AFM. The thickness of the films was studied by AFM and ellipsometry. The thicknesses of the Con A and the HRP film are about 9.0 and 4.6 nm, respectively, which correspond to their X-ray diffraction data. It is demonstrated that the shape of Con A and HRP molecule was well retained during the assembly process. Cyclic voltammetry (CV) was used to test the electrochemical properties of the films. The enzyme electrode of three bilayers of (Con A/HRP) films was sensitive for the reduction of hydrogen peroxide in the methylene blue (MB) solution. The response current is proportional to the concentration of H2O2 within the range of 0.2~1.0 mmol•L-1. The sensitivity of the enzyme electrode is 24.0 mA•mol-1 •L and the apparent Michaelis constant is 4.2 mmol•L-1.

Key words: HRP, Con A, Surface topography, Assembly film, AFM, Enzyme electrode