Excitation transfer in complexes of horse liver alcohol dehydrogenase

The protein fluorescence of LADH¹ was quenched upon coupling with NADH, NAD⁺, o-phenanthroline, or thyroxine and its related compounds, while AMP, ADP, ADPR, or NMN did not quench the fluorescence. Addition of isobutyramide or pyrazole to E₂R₂ or E₂O₂ did not alter the degree of quenching. The coupling of two molecules of NADH to one molecule of E₂I₂ caused an equal fluorescence enhancement for both molecules of NADH when excited in its 340-mμ absorption band. However, with excitation in the protein absorption range, it was found that the binding of the first NADH molecule to LADH caused a larger fluorescence change than the binding of the second one. This was ascertained by following the increase of the fluorescence caused by addition of excess E₂ to E₂I₂R₂, whereby the complexes E₂I₂R and E₂I₂ were formed. This seemed to indicate that excitation energy could be transferred from one subunit to the other in the same LADH molecule.