Time-resolved fluorescence spectroscopy based evaluation of stability of glucose oxidase.

Glucose oxidase (GOx) is one of the most frequently used enzymes in a design of enzymatic biosensors and biofuel cells, which are novel electrical energy generation systems. Therefore, a better understanding of the mode of action of this enzyme is very important for further development of GOx-based sensors. In this research fluorescence properties of GOx in different acidic media have been estimated by the evaluation of redox states of active center that is flavine adenine dinucleotide (FAD). Steady-state fluorescence spectroscopy was applied to monitor the activity of GOx. A variation of pH has been invoked to gain a better understanding in the variations of GOx activity. The tendency of GOx activity to decrease over the time was determined, while increased intensity of the fluorescence band of GOx at 530 nm was associated with a decreased activity of the enzyme. The changes in fluorescence intensity of this band are caused by the dissociation of FAD from the enzyme. This process is not reversible, therefore, the decrease in the fluorescence intensity can be also associated with structural changes of the FAD during its reduction. Unlabelled Image • Fluorescence properties of glucose oxidase (GOx) in different acidic medium have been estimated. • Steady-state spectroscopy was applied to monitor the activity of glucose oxidase. • Evaluation of redox states of active center – FAD – has been performed. • The tendency of GOx activity decrease over the time was determined. • The changes of fluorescence intensity band have been associated with dissociation of FAD from the protein pat of enzyme. [ABSTRACT FROM AUTHOR]