Structure and properties of 5-deazaflavin radicals as compared to natural flavosemiquinones.

In order to gain more insight into flavin radicals, on which the selection of 1e−- and 2e−-oxireduction modes in flavoproteins depends, we have investigated structure, spectral properties and decay mode of molecular species occurring in the half-reduced 5-deazaflavin ‘model’ system by flash photolysis and pulse radiolysis. 1. (1) Enforced 1e−-reduction of 5-deazaflavin yields the short-lived red-colored 1-HdḞl, which is a strong reductant. In the absence of any electron acceptor, this radical decays by 1,5-prototropy (see below) and dismutation, which is rapidly reversed upon illumination. Competing with this photo-comproportionation, irreversible formation of the photo-stable σ-dimer (HdFl)2, linked via C(5), is observed, which becomes prevalent under prolonged illumination. 2. (2) Enforced 1e−-abstraction from 1,5-dihydro-5-deazaflavin yields the tautomeric 5-HdḞl, which is a mild oxidant and is transparent at λ > 480 nm. Prototropy 5-HdFl ⇆ 1-HdFl can be rate-determining in 5-deazaflavin redox reactions. Hence, the radical state in the 5-deazaflavin system does not mediate double 1e−-oxidoreduction as do natural flavosemiquinones. Instead, 5-deazaflavin favors nucleophilic substrate addition (carbanion transfer) and formation of intermediate σ-adducts in (photo)reductions even over the extent obsersed with natural flavin. This confirms the description of 5-deazaflavin as a ‘flavin-shaped nicotinamide derivative’. It explains at the same time the mechanism of 5-deazaflavin acting as a mild and yet potent photosensitizer in 1e−-reductions of biological redox systems. 3. (3) It is shown that replacement of N(5) by CH in the flavin nucleus also leads to the disappearance of the known action-pK in the photoreduction, which confirms the assignment of the latter pK in the natural flavin system to 5-protonation of the excited flavin triplet. From these model studies the following biological conclusions can be confirmed: The tautomer equilibrium of natural flavin semiquinones is diffusion-controlled and regulated thermodynamically: 5HFl ⇄ 1-HFl, while in flavoproteins the same equilibrium is regulated by regiospecific H-bridges from the apoprotein, which thus decides between 1e−- (stable 5-HFl) and 2e−-reaction (unstable 1-HḞl) modes.