Complex formation and intermolecular electron transfer between flavocytochrome b2 in the crystal and cytochrome c.
The present study addresses the question whether tetrameric flavocytochrome b2 in the crystal is catalytically competent and, if so, whether it is possible to prepare a functional complex of the crystalline enzyme with its physiological electron acceptor cytochrome c. By single crystal microspectrophotometry we show that the native reduced enzyme can be oxidized by oxygen or ferricyanide and that the oxidized enzyme can be reduced by the electron donor L-lactate. Reduced cytochrome c appears to diffuse through the liquid channels of flavocytochrome b2 crystals and, at low ionic strength, to accumulate in amounts stoichiometrically equivalent to the enzyme protomers. Both cytochromes can be oxidized by ferricyanide. In the presence of L-lactate, both cytochromes become reduced. Since reduction of cytochrome c by L-lactate requires the catalytic action of flavocytochrome b2, it is concluded that the structure of the crystalline enzyme not only allows for electron transfer from L-lactate to flavin and intramolecular electron transfer from flavin to heme, but also for the formation of a productive complex with cytochrome c.
D003460:Crystallization; D003574:Cytochrome c Group / Q000378:metabolism*; D004579:Electron Transport; D005292:Ferricyanides / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism*; D042844:L-Lactate Dehydrogenase (Cytochrome); D010100:Oxygen / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013056:Spectrophotometry, Ultraviolet
D003574:Cytochrome c Group; D005292:Ferricyanides; C007931:hexacyanoferrate III; D007770:L-Lactate Dehydrogenase; D042844:L-Lactate Dehydrogenase (Cytochrome); D010100:Oxygen