Time-resolved fluorescence of the two tryptophans in transducin α-subunit

摘要

G{sub}α contains two tryptophans. Tryptophan fluorescence decay of G{sub}α at 25°C in 10 mM Tris-HCl buffer pH=7.3 was triexponential with lifetimes of 3.62, 1.06 and 0.17ns. Contrary to expectation, the shorter lived (lower quantum yield) components were blue-shifted relative to the longer Denaturation with 8M urea diminished the 1.06ns component and increased net intensity which became dominated by a lifetime of 5.10ns. Emission of a tryptophan species was identified at 450nm with lifetime of 20-40ns, similar to that reported earlier by Vanderkooi J. et al. This long-wavelength emission was also eliminated by 8M urea denaturation, making it seem possible that the low intensity of the 1.06ns blue shifted component may result from competing excited state processes that give rise to the 450nm emission. Transient absorbance spectra of G{sub}α show no evidence for triplet states in denatured protein while the native protein showed phosphorescence at ～442nm of lifetime between 0.1 and 1.0ms.