Time-Resolved Flavin Adenine Dinucleotide Fluorescence Study of the Interaction Between Immobilized Glucose Oxidase and Glucose

Time-resolved fluorescence experiments have shown that flavin adenine dinucleotide (FAD) fluorescence emission of sol-gel immobilized glucose oxidase (GOD) exhibits a three-exponential decaying behaviour characterized by long- (about 2.0-3.0 ns), intermediate- (about 300 ps) and short- (less than 10 ps) lifetime, each one being characteristic of a peculiar conformational state of the FAD structure. In the present work time-resolved fluorescence is used to monitor FAD signals in the time interval immediately following the addition of glucose at various concentrations in order to detect the conformational changes occurring during the interaction between sol-gel immobilized GOD and glucose. The analysis of time-dependent fluorescence emission signal has shown that the FAD conformational state changes during the process from a configuration with a prevalence of the state characterized by the long lifetime to a configuration with increased contribution from the process with the intermediate lifetime. The time needed to complete this configuration change decreases with the concentration of added glucose. The results here reported indicate that time-resoled fluorescence can be extremely useful for a better understanding of solid phase biocatalysis that is particularly important in light of their clinical and biotechnological applications.