RESONANCE RAMAN-STUDY ON THE OXIDIZED AND ANIONIC SEMIQUINONE FORMS OF FLAVOCYTOCHROME B(2) AND L-LACTATE MONOOXYGENASE - INFLUENCE OF THE STRUCTURE AND ENVIRONMENT OF THE ISOALLOXAZINE RING ON THE FLAVIN FUNCTION

The oxidized and semiquinone anion radical forms of flavin mononucleot ide carried by flavocytochrome b(2) and L-lactate monooxygenase have b een studied by resonance Raman (RR) spectroscopy, The RR spectra of th eir oxidized forms are compared with previously published RR data on v arious flavins and flavoproteins. Taking as a support available X-ray crystallographic data on flavoproteins, we have found correlations bet ween the frequencies of RR bands II (1575-1588 cm(-1)), III (1534-1557 cm(-1)), and X (1244-1266 cm(-1)) and the H-bonding environment and/o r the structure of the flavin ring. The present RR data provide strong evidence that the electron density, the conformation, and tile H-bond ing environment of the oxidized flavin mononucleotide of flavocytochro me b(2) and L-lactate monooxygenase are different, As far as the anion ic semiquinone form of flavoproteins is concerned, the behavior of two bands observed at 1280-1300 and 1320-1350 cm(-1) suggests that they h ave vibrational origins similar to those of RR bands II and III of oxi dized compounds, On this basis, the differences in conformation and B- bonding environment of the isoalloxazine ring, observed for the oxidiz ed form of flavocytochrome b(2) and L-lactate monooxygenase, appear to be preserved upon one-electron reduction of the flavin, For both flav oproteins, the RR spectra of the semiquinone form are affected by pyru vate binding, The data are interpreted in the frame of a change in II- bonding interaction of the C4=O carbonyl group of the flavin without s ignificant alteration of the isoalloxazine conformation, This modifica tion in electrostatic interaction quantitatively accounts for the pyru vate-induced changes of the oxidized/semiquinone and semiquinone/reduc ed redox potentials of the flavoproteins. Considering the high homolog y in the flavin catalytic sites of flavocytochrome b(2) and L-lactate monooxygenase, the observed differences in Pi-bonding environment and conformation of the FMN ring are related to the different biological f unctions of the two flavoproteins.