Abstract
Flavocoenzymes are extremely versatile but the reaction catalyzed by each individual flavoenzyme is highly specific. This requires the prerequisite mechanism that controls the flavin reactivity. In order to understand this control mechanism, the author has investigated various flavoenzymes focusing on how the flavin reactivity is controlled by monitoring the electronic state of the flavin moiety and its fluctuation along the reaction pathway. In the present report, two flavoenzymes, _D-amino acid oxidase and medium-chain acyl-CoA dehydrogenase, are concerned, the major method being ^<13>C- and ^<15>N-NMR spectroscopy of these enzymes reconstituted with ^<13>C- and ^<15>N-enriched FAD. The results obtained indicate the following : 1. The reactivity of flavin in the oxidized form of the enzymes is controlled by hydrogen-bonding network in the protein matrix and by its fluctuation during the reaction sequence. 2. In the control of the flavin reactivity, the overlap between LUMO (lowest unoccupied molecular orbital) of flavin and HOMO (highest occupied molecular orbital) of the substrate plays an important role. 3. The reoxidation process of the reduced enzymes is controlled by modulation of the electron density at C (4a) of reduced flavin, this position being the site of nucleophilic attack to molecular oxygen.Flavocoenzymes are extremely versatile but the reaction catalyzed by each individual flavoenzyme is highly specific. This requires the prerequisite mechanism that controls the flavin reactivity. In order to understand this control mechanism, the author has investigated various flavoenzymes focusing on how the flavin reactivity is controlled by monitoring the electronic state of the flavin moiety and its fluctuation along the reaction pathway. In the present report, two flavoenzymes, _D-amino acid oxidase and medium-chain acyl-CoA dehydrogenase, are concerned, the major method being ^<13>C- and ^<15>N-NMR spectroscopy of these enzymes reconstituted with ^<13>C- and ^<15>N-enriched FAD. The results obtained indicate the following : 1. The reactivity of flavin in the oxidized form of the enzymes is controlled by hydrogen-bonding network in the protein matrix and by its fluctuation during the reaction sequence. 2. In the control of the flavin reactivity, the overlap between LUMO (lowest unoccupied molecular orbital) of flavin and HOMO (highest occupied molecular orbital) of the substrate plays an important role. 3. The reoxidation process of the reduced enzymes is controlled by modulation of the electron density at C (4a) of reduced flavin, this position being the site of nucleophilic attack to molecular oxygen.
