Homogeneous Spectrally- and Time-Resolved Fluorescence Emission from Single-Tryptophan Mutants of IIA^Glc Protein

Abstract

We report an experimental study of protein relaxation dynamics on the picosecond and nanosecond time scales. The protein equilibrium state is perturbed by the redistribution of electric charge density over the side chain of a tryptophan residue. Electronic excitation of the residue induces the charge shift and triggers a relaxation process, the dynamics of which is reflected in tryptophan fluorescence emission. In the case of homogeneous emission, the relaxation dynamics can be extracted from a time-dependent red shift in the emission spectrum. In the case of heterogeneous emission, the spectral shift may not represent relaxation dynamics. A criterion for distinguishing between homogeneous and heterogeneous fluorescence emission is suggested here. Emission from the mutants E21W and F3W of IIA Glc is found to be free from permanent or long-lived heterogeneity. In E21W, the only tryptophan residue is in a rigid globule, whereas in F3W it is on a flexible tail. The relaxation dynamics reported by the tryptophan residue in F3W is much faster than that in E21W. Addition of glycerol to the solvent slows down the relaxation dynamics for both tryptophan residues.

Resolving environmental microheterogeneity and dielectric relaxation in fluorescence kinetics of protein

“…More recently descriptions combining the conformational heterogeneity (rotamers) with dielectric relaxation have emerged [9][10][11][12][13][14][15][16]. In the original paper by Toptygin and Brand [9], the spectrally-and time-resolved fluorescence emission I  (,t) [9][10][11][12], is written as …”

“…In the original paper by Toptygin and Brand [9], the spectrally-and time-resolved fluorescence emission I  (,t) [9][10][11][12], is written as …”

Fluorescence quenching of tryptophan and tryptophanyl dipeptides in solution

“…As a result, the effect of the environment on the fluorescence maxima and spectral width is fairly well understood. This is the case even though environment effects can tune the fluorescence maxima of tryptophan in proteins to values within a 310 -350 nm range, quantum yields from virtually 0 to 0.35, and fluorescence lifetimes from 50 to 8000 ps [2,[10][11]. But until recently, there has been no good model to relate quantum yield to the nature of the tryptophan microenvironment.…”

Chapter 8 Ultrafast Fluorescence Spectroscopy via Upconversion

“…Neither wavelength region exhibited terms slower than 2 ps, however, except for previously measured terms well over 50 ps (e.g., in the TCSPC work of Toptygin and Brand) (Toptygin et al, 2001). The exposed location of Trp in the mutant E21W might lead us to think of it as neat Trp.…”

“…Generalized solvent relaxation yields short-lived blue and long-lived red DAS on the nanosecond timescale (Alcala et al, 1987;Toptygin et al, 2001;Vincent et al, 1995).…”