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Keywords:

multifrequency phase fluorometry, electron-transfer reactions, protein interactions, indole fluorescence, escherichia-coli, dsba protein, peptide-bond, side-chains, decay, trichosanthin, Science & Technology, Life Sciences & Biomedicine, Biophysics, MULTIFREQUENCY PHASE FLUOROMETRY, ELECTRON-TRANSFER REACTIONS, PROTEIN INTERACTIONS, INDOLE FLUORESCENCE, ESCHERICHIA-COLI, DSBA PROTEIN, PEPTIDE-BOND, SIDE-CHAINS, DECAY, TRICHOSANTHIN, Acrylamide, Algorithms, Crystallography, X-Ray, Dose-Response Relationship, Drug, Electrons, Fluorescence, Hydrogen, Kinetics, Models, Statistical, Molecular Conformation, Mutation, Protein Conformation, Spectrometry, Fluorescence, Time Factors, Trichosanthes, Tryptophan, 02 Physical Sciences, 03 Chemical Sciences, 06 Biological Sciences, 31 Biological sciences, 34 Chemical sciences, 51 Physical sciences

Abstract:

The Dead-End Elimination method was used to identify 40 low energy microconformations of 16 tryptophan residues in eight proteins. Single Trp-mutants of these proteins all show a double- or triple-exponential fluorescence decay. For ten of these lifetimes the corresponding rotameric state could be identified by comparing the bimolecular acrylamide quenching constant (k(q)) and the relative solvent exposure of the side chain in that microstate. In the absence of any identifiable quencher, the origin of the lifetime heterogeneity is interpreted in terms of the electron transfer process from the indole Cepsilon3 atom to the carbonyl carbon of the peptide bond. Therefore it is expected that a shorter [Cepsilon3-C=O] distance leads to a shorter lifetime as observed for these ten rotamers. Applying the same rule to the other 30 lifetimes, a link with their corresponding rotameric state could also be made. In agreement with the theory of Marcus and Sutin, the nonradiative rate constant shows an exponential relationship with the [Cepsilon3-C=O] distance for the 40 datapoints.