Flavin adenine dinucleotide, dissociation

The transient absorption spectra similar to that of the ion-pair state of indole cation radical and flavin anion radical were also observed in D-amino acid oxidase (5), although the spectra were not so clear as those of flavodoxin. In D-amino acid oxidase, the coenzyme, flavin adenine dinucleotide (FAD), is wealtly fluorescent. The fluorescence lifetime was reported to be 40 ps (16), which became drastically shorter (less than 5 ps) when benzoate, a competitive inhibitor, was combined with the enzyme at FAD binding site (17). The dissociation constant of FAD was also marlcedly decreased by the binding of benzoate (17). These results suggest that interaction between isoalloxazine and the quencher became stronger as the inhibitor combined with the enzyme. Absorbance of the transient spectra of D-amino acid oxidase-benzoate complex was remarkably decreased. In this case both rate constants of formation and decay of the CT state could become much faster than those in the case of D-amino acid oxidase free from benzoate. 

It is frequently the case that enzymes undergo slow heterogeneous electron transfer with an electrode surface. Such slow rates of electron transfer can largely be attributed to insulation of the redox active site by the bulk of the protein matrfac. This is especially true for en mes that have a non-dissociable redox cofactor, such as the flavin adenine dinucleotide (FAD) within the active site of glucose oxidase. In cases where an enzyme active site is physically inaccessible to the electrode surface, an artificial intermediate redox species is used to shuttle electrons between the enzyme and electrode. It should be noted at the outset that much of the early research in mediated bioelectrocatalysis was performed on homogeneous mixtures of substrate, enzyme and mediator. However, the vast majority of literature since the late 1980s has dealt exclusively with systems in which the enzyme and mediator are immobilized in some capacity at the electrode surface. Thus the majority of the topies eovered here are focused on immobilized systems to reflect the dominant trend in bioeleetrocatalysis. 

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