Intrinsic fluorescence spectra of a tryptophan-containing parvalbumin as a function of thermal, pH and urea denaturation.

Abstract

The thermal, pH and urea denaturation of the calcium-loaded protein from whiting has been studied by means of the intrinsic fluorescence of the single tryptophan residue. pH denaturation of the protein takes place at a pH greater than 11.5 and lower than 5.5. Thermal denaturation of the protein occurs at temperatures above 55 degrees C. Urea initiates the denaturation of the calcium-loaded protein at rather low concentrations (1.0 M). In all cases, whether pH, thermal or urea denaturation, intermediate states of the protein were recorded. The fluorescence spectra of these intermediates are similar to that of the protein with one equivalent of calcium bound. Whiting parvalbumin binds calcium in the presence of 7.5 M urea but under these conditions, calcium-binding constants of the protein have been shown to be 10(2)-10(3)M(-1) (in comparison with 5 X 10 (8) and 6 X 10(6)M(-1) in the absence of any denaturing agents).