Enzyme Surface Potential And Its Function Correlation Studies

Enzymatic reaction mechanism has been the hot spot in the life science research. Studies have confirmed that the conformation of enzymatic molecular changed after the enzymatic reaction, however, the dynamic change of enzyme molecules during the reaction has not been studied clearly. Zeta potential can effectively response enzyme molecular surface electric charge, thus reflect the change of enzyme molecular surface electrical. Therefore, this paper selects Zeta potential measurement indicators, combined with Infrared Spectroscopy and Circular Dichroism Spectroscopy, to explore the correlation of Zeta potential and the function of enzymes, and the change of the enzyme molecular properties in the process of the enzymatic reaction.The experiment results are as follows:The Zeta potential measurement of the enzyme molecule found that the Zeta potential of the enzyme molecules and substrate were affected by the measuring p H, temperature and ionic strength. After contacted with the substrate, the potential of the enzyme molecules changed to the electrically neutral direction rapidly. After the reaction, the potential of enzyme molecule gradually return to the direction of pre-reaction, but could not be restored to the state before contacting the substrate. The variation of Zeta potential increased with the increasing reaction temperature and adding amount of substrates. Took the α-amylase reactioned with starch dialyzed and lyophilized to obtain the induced α-amylase. After the starch was added to the enzyme solution its potential(absolute value) was further reduced. The α-amylase reactioned with starch and α-amylase was used for infrared spectrum, found similar IR spectra before and after reaction of the enzyme molecules. The Circular Dichroism Spectroscopy experiments suggested that the secondary structure changed rapidly after contacted with the substrate. After the reaction, the secondary structure of enzyme molecule gradually return to the direction of pre-reaction, but could not be restored to the state before contacting the substrate. And α-helix and β-turn showed the opposite trendency, which was associated with the Zeta potential change. Stopped flow experiment shown that the secondary structure of α-amylase have changed in 30 ms after contacted with the starch. Ion chromatography experiment suggested that the α-amylase has a small amount of product residue after reaction, but not enough to affect the potential measurement result. The heat resistance experiments of α-amylase showed that the change of temperature would lead to the potential and conformational changes of enzyme, and the presence of a substrate enhanced the thermal stability of α- amylase.At present, the research on the mechanism of enzymatic reaction more concentrated in the dynamic simulation and dynamics research, this paper studies the change of surface potential and secondary structure during the enzymatic reaction process, and explored the influence factors of the change. The experiment proved that the charge redistribution of enzyme molecule occurred during the enzymatic reaction, this echoes the induced fit theory, providing a new method for the study of the enzymatic reaction.