title: 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.
abstract: 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.
authors: Laboratory of Biomolecular Dynamics, Department of Chemistry, University of Leuven, Celestijnenlaan 200 G, B-3001 Leuven, Belgium.
authors: Giovanni
authors: G
authors: Maglia
authors: Abel
authors: A
authors: Jonckheer
authors: Marc
authors: M
authors: De Maeyer
authors: Jean-Marie
authors: JM
authors: Frère
authors: Yves
authors: Y
authors: Engelborghs
mesh_terms: D002268:Carboxypeptidases / Q000737:chemistry*; D005454:Fluorescence Polarization; D007700:Kinetics; D050505:Mutant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013302:Streptomyces / Q000201:enzymology*; D013816:Thermodynamics; D014364:Tryptophan / Q000737:chemistry*
chemical_list: D050505:Mutant Proteins; D014364:Tryptophan; D002268:Carboxypeptidases