Decomposition of Protein Tryptophan Fluorescence Spectra into Log-Normal Components. II. The Statistical Proof of Discreteness of Tryptophan Classes in Proteins

Abstract

The physical causes for wide variation of Stokes shift values in emission spectra of tryptophan fluorophores in proteins have been proposed in the model of discrete states (Burstein, E. A., N. S. Vedenkina, and M. N. Ivkova. 1973. Photochem. Photobiol. 18:263-279; Burstein, E. A. 1977a. Intrinsic Protein Luminescence (The Nature and Application). In Advances in Science and Technology (Itogi Nauki i Tekhniki), Biophysics Vol. 7. VINITI, Moscow [In Russian]; Burstein, E. A. 1983. Molecular Biology (Moscow) 17:455-467 [In Russian; English translation]). It was assumed that the existence of the five most probable spectral classes of emitting tryptophan residues and differences among the classes were analyzed in terms of various combinations of specific and universal interactions of excited fluorophores with their environment. The development of stable algorithms of decomposition of tryptophan fluorescence spectra into log-normal components gave us an opportunity to apply two mathematically different algorithms, SImple fitting with Mean-Square criterion (SIMS) and PHase-plot-based REsolving with Quenchers (PHREQ) for the decomposition of a representative set of emission spectra of proteins. Here we present the results of decomposition of tryptophan emission spectra of >100 different proteins, some in various structural states (native and denatured, in complexes with ions or organic ligands, in various pH-induced conformations, etc.). Analysis of the histograms of occurrence of >300 spectral log-normal components with various maximum positions confirmed the statistical discreteness of several states of emitting tryptophan fluorophores in proteins.

Thermostability of cardosin A from Cynara cardunculus L.

“…In titration experiments, pH values were adjusted by means of a polyethylene rod moistened with either 0.1 M HCl or 0.1 M NaOH. The fluorescence spectra were analysed on the basis of the model of discrete states of tryptophan in proteins [18][19][20].…”

The protein fluorescence and structural toolkit: Database and programs for the analysis of protein fluorescence and structural data

“…PFAST contains three modules: (1) FCAT is a fluorescence-correlation analysis tool, which decomposes protein fluorescence spectra to reveal the spectral components of individual tryptophan residues or groups of tryptophan residues located close to each other, and assigns spectral components to one of five previously established spectral-structural classes. (2) SCAT is a structural-correlation analysis tool for the calculation of the structural parameters of the environment of tryptophan residues from the atomic structures of the proteins from the Protein Data Bank (PDB), and for the assignment of tryptophan residues to one of five spectral-structural classes. (3) The last module is a PFAST database that contains protein fluorescence and structural data obtained from results of the FCAT and SCAT analyses.…”

Non-invasive characterization of structure and morphology of silk fibroin biomaterials using non-linear microscopy

“…Spectral shifts in response to a more polar environment have been observed for chromophores such as tryptophan, and are typically attributed to changes in the relaxation of the molecule because of polar interactions or the formation of hydrogen bound complexes in the excited state [48][49][50]. We expect that similar processes are responsible for the observed red shift in the silk spectra of samples exposed to more highly hydrated environments, depicted schematically in Fig.…”