Flavin and heme structures in lactate:cytochrome c oxidoreductase: a resonance Raman study.
Resonance Raman spectra of Hansenula anomala L-lactate:cytochrome c oxidoreductase (or flavocytochrome b2), of its cytochrome b2 core, and of a bis(imidazole) iron-protoporphyrin complex were obtained at the Soret preresonance from the oxidized and reduced forms. Raman contributions from both the isoalloxazine ring of flavin mononucleotide (FMN) and the heme b2 were observed in the spectra of oxidized flavocytochrome b2. Raman diagrams showing frequency differences of selected FMN modes between aqueous and proteic environments were drawn for various flavoproteins. These diagrams were closely similar for flavocytochrome b2 and for flavodoxins. This showed that the FMN structure must be very similar in both types of proteins, despite their very different proteic pockets. However, the electron density at this macrocycle was found to be higher in flavocytochrome b2 than in these electron transferases. No significant difference was observed between the heme structures in flavocytochrome b2 and in cytochrome b2 core. The porphyrin center-N(pyrrole) distances in the oxidized and reduced heme b2 were estimated to be 1.990 and 2.022 A from frequencies of porphyrin skeletal modes, respectively. The frequency of the vinyl stretching mode of protoporphyrin was found to be very affected in resonance Raman spectra of flavocytochrome b2 and of cytochrome b2 core (1634-1636 cm-1) relative to those observed in the spectra of iron-protoporphyrin [bis(imidazole)] complexes (1620 cm-1). These specificities were interpreted as reflecting a near coplanarity of the vinyl groups of heme b2 with the pyrrole rings to which they are attached. The low-frequency regions of resonance Raman indicated that the iron atoms of the four hemes b2 are in the porphyrin plane whatever their oxidation state. The histidine-Fe-histidine symmetric stretching mode was located at 205 cm-1 in the spectra of flavocytochrome b2 and of cytochrome b2 core. It was insensitive to the iron oxidation state and indicated strong Fe-His bonds in both states.
Desbois
A
A
Laboratoire de Biophysique, Institut de Biologie Physico-Chimique, Paris, France.
Tegoni
M
M
Gervais
M
M
Lutz
M
M
D005486:Flavin Mononucleotide / Q000032:analysis*; D006418:Heme / Q000032:analysis*; D007770:L-Lactate Dehydrogenase / Q000032:analysis*; D042844:L-Lactate Dehydrogenase (Cytochrome); D010084:Oxidation-Reduction; D011524:Protoporphyrins / Q000032:analysis; D013059:Spectrum Analysis, Raman
D011524:Protoporphyrins; D006418:Heme; D005486:Flavin Mononucleotide; D007770:L-Lactate Dehydrogenase; D042844:L-Lactate Dehydrogenase (Cytochrome)