摘要
建立了反相高效液相色谱法动态监测牛胰岛素在二硫苏糖醇存在下去折叠的过程。牛胰岛素在二硫苏糖醇作用下，首先发生构象变化，形成稳定的中间体后进一步断裂分子间的二硫键，形成Ａ链和Ｂ链。去折叠过程通过基质辅助激光解吸附质谱得到了鉴定。
A method for measurement of the dynamic unfolding procedure of bovine insulin by reversedphase HPLC has been established. Insulin contains 51 amino acids and two intrachain disulfide bridges. The denaturation of bovine insulin was carried in dithiothreitol solution at 100℃, and the equilibrium products were examined by HPLC at different reaction time. The results show that the conformation of insulin has changed before cleavage of the disulfide bonds to A and B chains. Bovine insulin, two intermediates and the reduction products A and B chains were well separated on a C 18 column (4.6mm×150mm) with a linear gradient elution of acetonitrile containing 0.1% trifluoroacetic acid. The conformation of the unfolding intermediates of insulin was indicated by chromatographic method, and the results were verified by matrixassisted laser desorption ionization time of flight mass spectrometry. The method is helpful to reveal the conformation changes in the procedures of protein unfolding.
出处
《色谱》
CAS
CSCD
北大核心
1997年第5期420-422,共3页
Chinese Journal of Chromatography
关键词
高效液相色谱
牛胰岛素
二硫苏糖醇
去折叠
reversedphase high performance liquid chromatography, MALDITOF, bovine insulin, dithiothreitol, unfolding
