Premium
An unusual red‐edge excitation and time‐dependent Stokes shift in the single tryptophan mutant protein DD‐carboxypeptidase from Streptomyces : The role of dynamics and tryptophan rotamers
Author(s) -
Maglia Giovanni,
Jonckheer Abel,
De Maeyer Marc,
Frère JeanMarie,
Engelborghs Yves
Publication year - 2008
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.073147608
Subject(s) - tryptophan , conformational isomerism , chemistry , fluorescence , excitation , stokes shift , protein dynamics , molecular dynamics , biophysics , crystallography , chemical physics , computational chemistry , molecule , physics , amino acid , biochemistry , biology , optics , organic chemistry , quantum mechanics
The fluorescence emission of the single tryptophan (W233) of the mutant protein DD‐carboxypeptidase from streptomyces is characterized by a red‐edge excitation shift (REES), i.e., the phenomenon that the wavelength of maximum emission depends on the excitation wavelength. This phenomenon is an indication for a strongly reduced dynamic environment of the single tryptophan, which has a very low accessibility to the solvent. The REES shows, however, an unusual temperature and time dependence. This, together with the fluorescence lifetime analysis, showing three resolvable lifetimes, can be explained by the presence of three rotameric states that can be identified using the Dead‐End Elimination method. The three individual lifetimes increase with increasing emission wavelength, indicating the presence of restricted protein dynamics within the rotameric states. This is confirmed by time‐resolved anisotropy measurements that show dynamics within the rotamers but not among the rotamers. The global picture is that of a protein with a single buried tryptophan showing strongly restricted dynamics within three distinct rotameric states with different emission spectra and an anisotropic environment.