重组APP-N端蛋白与神经节苷脂GM1相互作用后的构象变化  

The Effects of GM1 on Conformational Changes of The N-terminus of APP

作　　者：丁吉新[1] 沙印林[2] 阮燕[1] 朱忠军[1] 黄力新[2] 耿慧敏[2] 聂松青[2] 张岱[1] 

机构地区：[1]北京大学精神卫生研究所,北京100083 [2]北京大学医学部生物物理系,北京100083

出　　处：《生物化学与生物物理进展》2003年第1期107-111,共5页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金资助项目 ( 39870 2 76;39840 0 10 );国家重点基础研究发展规划项目 (G19990 6 40 0 7) .~~

摘　　要：用基因重组方法制备人类 β 淀粉样蛋白前体 (APP)N端融合蛋白APP2 8～ 12 3 ,应用荧光光谱和圆二色谱技术观测GM 1对APP2 8～ 12 3 构象的影响 .结果表明 :APP2 8～ 12 3 与GM1水溶液和含GM1的脂质体孵育后 ,其荧光强度明显增强 ,最大发射峰位蓝移 2 0nm ;APP2 8～ 12 3 在溶液中的二级结构以α螺旋为主 ,峰位在 2 0 8nm和2 2 2nm ,而APP2 8～ 12 3 与PC/GM 1脂质体或GM 1水溶液孵育后 ,其二级结构虽以α螺旋为主 ,但摩尔椭圆度(θ)值明显增强 .以上结果表明GM1改变了APP2 8～ 12 3 二级结构 ,提示GM1引起APP分子构象的改变 ,可能影响APP分子正常生理功能和跨膜转运过程 .Alzheimer's disease (AD) is neuropathologically characterized by the presence of extracellular amyloid plaques and intracellular neurofibrillary tangles. The core of senile plaque is amyloid beta-protein (Abeta), which comes from its precursor - amyloid beta-protein precursor (APP). The pQE-APP(28similar to123) plasmids was constructed by gene recombination. The APP(28similar to123) protein was expressed in Escherichia coli and then purified. The purified products were examined for GM1 binding abilities by Western blotting. The effects of GM1 on conformation of APP N-terminus were detected by fluorescence and circular dichroism (CD) techniques. APP(28similar to123) protein could bind with GM1. The intrinsic fluorescence intensity of APP(28similar to123) protein in PC/GM1 vesicles or GM1 solution remarkably increased and the fluorescence peak value blue shifted 20 nm. CD results showed that the major secondary structure of APP(28similar to123) in PBS buffer was alpha-helix. When APP(28similar to123) incubated with PC/GM1 vesicles or GM1 solution, the alpha-helix content increased markedly. These results suggested that GM1 might affect the physiological function of APP and change APP span-membrane process and interfere APP trafficking and internalization by anchoring this molecule on the membrane, which may provide much more substrate for gamma-secretase and enhance Abeta generation on the cells.

关 键 词：重组APP-N端蛋白 神经节苷脂 GM1 相互作用 构象变化 Β-淀粉样蛋白前体 阿尔茨海默病 

分 类 号：R749.16[医药卫生—神经病学与精神病学]