Conformation heterogeneity in proteins as an origin of heterogeneous fluorescence decays, illustrated by native and denatured ribonuclease T₁

We examined the frequency-domain intensity decays of the intrinsic tryptophan fluorescence (Trp-59) from ribonuclease T₁ (EC 3.1.27.3) (RNAase T₁). At pH 5.5 in the native state (below 30°C), the intensity decay of the single tryptophan residue is a single-exponential process. Conditions which result in protein unfolding were found to induce more complex intensity decays. At temperatures above 40°C, or in the presence of guanidine hydrochloride, the intensity decays became obviously double exponential. In general, the main effect of temperature or guanidine was to induce a second subnanosecond component in the intensity decay. The increased complexity of the decays could not be explained by a unimodal distribution of decay times. These results indicate that conformational dispersion of protein structure can be one origin of the multi-exponential decays which are generally observed for protein fluorescence.