FLUORESCENCE AND THE LOCATION OF TRYPTOPHAN RESIDUES IN PROTEIN MOLECULES

Abstract

Abstract— Fluorescence spectra of a number of native and denaturated proteins have been analysed, using spectral band width (ΔΛ), spectral maximum position (Λ m ), fluorescence quenching by external ionic quenchers, lifetime (b), and quantum yield ( q ) and its changes upon denaturation. The results enabled a model of fluorescence properties of tryptophan residues in the proteins to be substantiated by considering the existence of three discrete spectral classes, one buried in nonpolar regions of the protein (Λ m 330–332 nm, ΔΛ= 48–49nm, q 0.11, τ= 2.1 ns) and two on the surface. One of the latter is completely exposed to water (Λ m # 350–353 nm, ΔΛ= 59–61 nm, q # 0.2, τ= 5.4 ns); the other is in limited contact with water which is probably immobilized by bonding at the macromolecular surface (Λ m # 340–342 nm, ΔΛ= 53–55 nm, q # 0.3, = 4.4 ns). Some quantitative predictions from the model, for (a) the fraction of fluorescence that is quenched by ionic quenchers, (b) the mean values of quantum yield, and (c) the mean values of fluorescence lifetime for various proteins, show good concordance with independent experimentally determined values.