Spectroscopic evidence for NADH-induced conformational changes in rabbit muscle aldolase.

The intrinsic fluorescence (steady-state spectra, anisotropy and nanosecond decay) in combination with phosphorescence at room temperature were used to detect and characterize conformational changes in rabbit muscle aldolase accompanying the NADH-binding process. Ligand binding has entailed a decrease in aldolase fluorescence intensity, a blue shift in its maximum and a polarization increase in a long wavelength part of the emission spectrum. The NADH binding induces the changes in room temperature phosphorescence - higher intensity and longer lifetime. The excited state energy transfer from tryptophans to NADH is not observed, and the character of spectroscopic changes on NADH binding allows us to reveal the spectroscopic heterogeneity among the tryptophan residues. The character, location of protein conformational changes associated with the binding of NADH and their relation to the tryptophans' microenvironment in aldolase are discussed.