L-Lactate cytochrome c reductase: rapid kinetic studies of electron transfers within the flavocytochrome b2-cytochrome c assembly.

This study is part of a series aimed at the characterization of individual steps of electron transfer taking place between prosthetic flavin, heme b2, heme c within active sites and complexes. After rapid mixing of ferricytochrome c with partially reduced flavocytochrome b2, the reaction is followed at the level of two reactants, cytochrome b2 and cytochrome c. In order to define the proper reactivity of flavosemiquinone, conditions under which this form is highly stabilized (presence of pyruvate) have been chosen. With the help of simulations, it has been possible to characterize a rapid step of electron transfer from cytochrome b2 to cytochrome c within a complex (at approx. 70% saturation) and a slow step k = 5 s-1 assigned to cytochrome b2 reduction by flavosemiquinone within the active site of the pyruvate-liganded enzyme.