Binding of Two Coenzyme Molecules

Although a thermodynamic and kinetic framework for electron transfer in CPR has emerged from rapid-mixing kinetic and potentiometry studies , alternative approaches have been required to access the rate of internal electron transfer between the flavins. Tollin and coworkers have used flash photolysis studies employing pho-toexcited 5-deazariboflavin to investigate the kinetics of inter-flavin electron transfer in rabbit CPR . CPR is reduced rapidly by photoexcited 

5-deazariboflavin (6.8 X lO M s ) and this phase is followed by a slower (70 s ) partial loss of blue semiquinone signal that reports on interflavin electron transfer from FAD to FMN. Pre-reduction of rabbit CPR prior to flash photolysis yields a rate constant for internal electron transfer of 15 s . This slower rate for pre-reduced enzyme (FAD, /FMN ) is consistent with the smaller driving force for electron transfer from FADjq to FMNjjj following electron donation by 

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