摘要
运用荧光光谱学手段研究了含有FMN的大肠杆菌偶氮还原酶AzoR以及R59G突变体与底物酸性红之间的相互作用,发现伴随底物酸性红浓度的增加,FMN在520nm左右的荧光强度显著降低。通过标准结合方程拟合获得了酶与底物的结合常数,发现25℃野生型AzoR与酸性红的结合常数为2.84×106 L·mol-1,而突变体R59G的则降到了4.75×105 L·mol-1;此外,通过对野生型及其突变体进行变温实验、并计算焓变和熵变,发现当第59位精氨酸突变成甘氨酸后会导致底物与酶之间的相互作用类型从离子键向范德华力转变。
In this work the interaction between FMN-containing AzoR and acid red has been investigated by utilizing fluorescence spectroscopic methods.It was observed that the fluorescence intensity of the cofactor FMN at 520 nm in AzoR decreased significantly upon increasing the concentration of AR.The associate constant was obtained using standard binding equation,and the resulting associate constant was found to decrease from 2.84×106 L·mol-1in wt-AzoR to 4.75×105 L·mol-1 in R59 Gat 25℃.In addition,we also performed the fluorescence quenching experiments at 37 ℃.The thermodynamic parametersΔH andΔS were calculated,and their symbol analysis showed that the 59 th arginine mutation into glycine can lead to the interaction between the substrate and enzyme types change from the electrostatic forces to van der Waals force.
出处
《光谱学与光谱分析》
SCIE
EI
CAS
CSCD
北大核心
2016年第S1期375-376,共2页
Spectroscopy and Spectral Analysis
