Reinvestigation of the structure of oxidized and reduced flavin: carbon-13 and nitrogen-15 nuclear magnetic resonance study.

Several chemically substituted flavins are investigated in the oxidized and the reduced state by 13C and 15N NMR techniques. The dependence on the polarity of the solvent and on the concentration is studied. In combination with already published results, a semiempirical theory is developed to interpret the chemical shifts in terms of the solution structure of flavins. Where possible, the results are compared with crystallographic and light absorption data. In contrast to common ideas, the solution structure of the oxidized state is not fully coplanar, but the N(10) atom is situated out of plane to a certain degree. Polarizing the flavin by hydrogen bonds in a high dielectric medium moves the N(10) atom into the molecular plane, and the flavin molecule becomes coplanar. In the coplanar molecule, pi electrons are delocalized from the N(10) atom mainly to O(2 alpha) and O(4 alpha). The NMR results show that the solution structure of reduced flavin is mainly governed by sterical hindrance and hydrogen bonds. The findings are in contrast to commonly accepted ideas that reduced flavin is strongly bent. In an apolar solvent, the reduced neutral isoalloxazine is only slightly bent. The formation of hydrogen bonds in a protic solvent of a high dielectric constant decreases the bend. The N(10) atom is now almost fully sp2 hybridized, and the N(5) atom has an endocyclic angle of 115-117 degrees, indicating its predominant sp2 character. The results have several important implications for flavin catalysis.(ABSTRACT TRUNCATED AT 250 WORDS)