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Flavoproteins are a class of enzymes catalyzing a very broad spectrum of redox processes by different chemical mechanisms. This review describes the best studied of these mechanisms and discusses factors possibly governing reactivity and specificity.A large number of flavin-containing enzymes, several hundreds, has been uncovered to date. An unusual feature of flavoproteins is the variety of the catalytic reactions performed, which range from typical redox catalysis such as the dehydrogenation of an amino acid, or the activation of dioxygen, to photochemistry; from 'DNA damage repair' to light emission. These few examples illustrate the fact that the same coenzyme is able to catalyze, or take part in catalytic events which must vary widely from a mechanistic point of view. The chemistry underlying the conversion itself will be quite different from case to case. This versatility sets flavoproteins apart from most other cofactor-dependent enzymes, which, in general, each catalyze a single type of chemical reaction. The activation or 'steering' of a particular activity of the flavin results from the interaction with the protein at the active center. On the other hand, for the vast majority of these enzymes a common feature exists, that at some stage during the catalytic event a transfer of electrons takes place between the substrate and the flavin itself.The purpose of the present review is not to enumerate the different functions of flavoproteins, this having been done elsewhere [l -41, but to focus on the mechanisms of catalysis, and on the possible ways of interaction of the flavin nucleus with the protein, i.e. on how this brings about chemistry appropriate to the particular enzyme reaction. In order toCorrespondence to S. Ghisla,

Since their discovery and chemical characterization in the 1930s, flavins have been recognized as being capable of both one- and two-electron transfer processes, and as playing a pivotal role in coupling the two-electron oxidation of most organic substrates to the one-electron transfers of the respiratory chain. In addition, they are now known as versatile compounds that can function as electrophiles and nucleophiles, with covalent intermediates of flavin and substrate frequently being involved in catalysis. Flavins are thought to contribute to oxidative stress through their ability to produce superoxide, but at the same time flavins are frequently involved in the reduction of hydroperoxides, products of oxygen-derived radical reactions. Flavoproteins play an important role in soil detoxification processes via the hydroxylation of many aromatic compounds, and a simple flavoprotein in liver microsomes catalyses many reactions similar to those carried out by cytochrome P450 enzymes. Flavins are involved in the production of light in bioluminescent bacteria, and are intimately connected with light-initiated reactions such as plant phototropism and nucleic acid repair processes. Recent reports also link them to programmed cell death. The chemical versatility of flavoproteins is clearly controlled by specific interactions with the proteins with which they are bound. One of the main thrusts of current research is to try to define the nature of these interactions, and to understand in chemical terms the various steps involved in catalysis by flavoprotein enzymes.

ABSTRACT:The fluorescence and absorption properties of a series of reduced flavoproteins have been measured and compared with the properties of suitable model compounds. Contrary to common belief, a number of reduced flavoproteins have been found to exhibit appreciable fluorescence emission with maxima in the range 500-530 nm. In keeping with common observation, the reduced model flavines are devoid of fluorescence in solution at room temperature, but show marked fluorescence emission in the range 476-512 nm at 77 OK in rigid glasses. The fluorescence quantum yield of reduced lactate oxidase (emission x , , , 507 nm) is increased 4.7 times upon formation of covalent N5 adducts and the emission maximum is shifted to 476 nm. In the case of the nonfluores-T he electronic properties of the isoalloxazine chromophore system, which constitutes the redox active moiety of the flavine coenzymes, have been the object of several thorough theoretical and experimental investigations (Sun et al., 1972 ;Song, 1971, and literature cited therein). These studies have dealt mainly with the oxidized form of the free and proteinbound flavine as well as with model compounds. A distinctive characteristic of oxidized free flavines is their relatively strong fluorescence (quantum yield -0.3; Sun et al., 1972), with an emission maximum typically around 520 nm. The energy and intensity of this emission are dependent on solvent polarity and temperature (Koziol, 1969; Sun el al., 1972;Kotaki et al., 1967), on formation of complexes with a variety of molecules (Weber, 1950; Slikin, 1971), and on the position and properties of substituents Walker et al., 1972; Sun et al., 1972). Similar effects have been observed with riboflavine, FMN, or FAD1 bound to various apoproteins.Unfortunately the spectroscopy of reduced flavines has received far less attention than that of the oxidized flavoquinones. A reason for this could be the absence of wellresolved structure in the near-ultraviolet absorption spectrum and of fluorescence at ambient temperature in any solvent.On the other hand, the nonplanarity of the flavine ring in its reduced forms introduces difficulties in theoretical calculations

This work was performed during tenure of Career Development Award GM-K3-31.213.1 Abbreviations used: epr, electron paramagnetic resonance;FMN, riboflavin 5'-phosphate; TPNH, reduced triphosphopyridine nucleotide; FAD, FADH, FADHi, oxidized, semiquinoid, and fully reduced flavin-adenine dinucleotide; TPN+, oxidized triphosphopyridine nucleotide.cals, which are very readily distinguished on the basis of their optical absorption properties. With one enzyme, glucose oxidase, both types of radical are found, the concentrations depending on the pH. These results suggest that the two different spectral species may be due to the neutral and anionic radical forms of the flavin coenzyme prosthetic groups.et al., 1966) and Azotobacter flavoprotein (Beinert, 1965). Again this long wavelength absorption has been found to be well correlated with an epr-detectable free radical, and experimental extinction coefficients in the range 3000-5000 1. mole-1 cm-1 have been found at 570 µ. In the case of two other flavoproteins, d-