Measurements of the Physicochemical Properties of Proteins by Capillary Electrophoresis

• Main Authors: Chi Chung Chih, 植啟中
• Other Authors: Chunhung Wu
• Format: Others
• Language: zh-TW
• Published: 2005
• Online Access: http://ndltd.ncl.edu.tw/handle/78560939569949282288

碩士 === 淡江大學 === 化學學系 === 93 === We have developed a novel analytical method combining Poiseuille’s Law, Taylor-Aris Dispersion method and capillary electrophoretic technique for the simultaneous measurements of buffer viscosity and protein diffusion coefficient and mobility. The hydrodynamic radius and the effective charge of protein in buffer solutions with different pH values and ionic strengths can thus be calculated by using this method. For the same protein in buffer solutions with pH values higher than 5.6, the measured protein sizes did not change significantly. For monomeric proteins with spherical shape, the calculated protein volumes had good linear relationship with their corresponding molecular weights. This implicated that the measured proteins had similar densities at that pH value. At pH values lower than 4.6, no linear relationship between protein volume and molecular size was found for the studied proteins, which we attributed to the different partially denatured conformations possessed by different proteins. At relatively low ionic strength it was found that the measured diffusion coefficient of protein increased with decreasing ionic strength, which was mainly due to the increase in electrostatic repulsion force among protein molecules. Under these conditions the value of diffusion coefficient was not only affected by the size of protein, but also affected by the electrostatic repulsion among protein molecules. Therefore, the hydrodynamic radius of protein could not be calculated directly by using the Stoke-Einstein equation. In this research, we discussed the binding behaviors between protein and sodium dodecylsulfate(SDS) by using the newly developed analytical method as described above. We found that the measured sizes and mobility of protein-SDS complexes were affected by protein property, the concentration of SDS, and the buffer solutions with different pH, ionic strength and viscosity.