Journal ArticleDOI
Exposure of tryptophanyl residues in proteins. Quantitative determination by fluorescence quenching studies.
TLDR
The value of this probing technique lies in its ability to sense not only the steady-state exposure of a residue in a protein, but also its dynamic exposure.Abstract:
Acrylamide is an efficient quencher of tryptophanyl fluorescence which we report to be very discriminating in sensing the degree of exposure of this residue in proteins. The quenching reaction involves physical contact between the quencher and an excited indole ring, and can be kinetically described in terms of a collisional and a static component. The rate constant for the collisional component is a kinetic measure of the exposure of a residue in a protein, and values ranging from 4 X 10(9) M-1 S-1 for the fully exposed tryptophan in the polypeptide, adrenocorticotropin, to less than 5 X 10(8) M-1 S-1 for the buried residue in azurin have been found. Static quenching is readily detected in proteins that are denatured, or contain only a single fluorophor. Quenching patterns for most multi-tryptophan containing proteins are difficult to analyze precisely, but qualitative information can, nevertheless, be extracted. Applications of this probing technique for monitoring protein conformational changes, such as the acid-induced expansion of human serum albumin, and inhibitor binding to enzymes, are presented. The value of this method lies in its ability to sense not only the steady-state exposure of a residue in a protein, but also its dynamic exposure.read more
Citations
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Journal ArticleDOI
Roles and Microenvironments of Tryptophanyl Residues of Spinach Phosphoribulokinase
TL;DR: Kinetic parameters of the W241F mutant are not significantly altered relative to those of wild-type enzyme, thereby discounting any requirement for Trp at position 241, and substitution of Trp155 with Phe or Ala has little impact on Vmax, but the Km for Ru5P and ATP are increased substantially; the diminished affinity for ATP is particularly pronounced in the case of the Ala substitution.
Journal ArticleDOI
Negative Cooperativity and High Affinity in Chitooligosaccharide Binding by a Mycobacterium smegmatis Protein Containing LysM and Lectin Domains.
TL;DR: Interestingly, in the case presented here, two LysM domains are enough for binding to peptidoglycan in contrast to the three reportedly required by the LysM domain of Bacillus subtilis and Lactococcus lactis.
Journal ArticleDOI
Structural and functional differences between fetal and adult serum albumin
TL;DR: The findings suggest that fetal and adult human serum albumin have a very similar three-dimensional structure; the different binding capacity for bilirubin displayed by the two proteins is likely to be the consequence of small differences in the physico-chemical properties of some amino acid residues close to the bilirUBin binding site.
Journal ArticleDOI
Indole to phenolate energy transfer, the alkaline quenching mechanism of β-trypsin fluorescence
TL;DR: Evidence that indole to phenolate energy transfer causes the decrease in beta-trypsin fluorescence with increasing alkalinity is obtained indirectly from viscosity, lifetime, wavelength dependence and chemical modification studies which either exclude or minimize the involvement of other possible mechanisms.
Journal ArticleDOI
A tryptophan residue is identified in the substrate binding of penicillin G acylase from Kluyvera citrophila
TL;DR: The penicillin G acylase purified from Kluyvera citrophila (KcPGA) on modification with tryptophan-specific reagents such as N-bromo succinamide (NBS) and 2-hydroxy 5-nitrobenzylbromide (HNBB) showed partial loss of activity and substrate protection.
References
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Journal ArticleDOI
The interpretation of protein structures: estimation of static accessibility.
B. Lee,Frederic M. Richards +1 more
TL;DR: The accessibility of atoms in the twenty common amino acids in model tripeptides of the type Ala-X-Ala are given for defined conformation and the larger non-polar amino acids tend to be more “buried” in the native form of all three proteins.
Journal ArticleDOI
Removal of Fatty Acids from Serum Albumin by Charcoal Treatment
TL;DR: Fluorescence spectra of human serum albumin samples indicated that impurities are sometimes present which can be removed by charcoal at neutral pH, and acid-charcoal treatment is a much more rapid method of removing lipid impurities than other methods previously described.
Journal ArticleDOI
Solute perturbation of protein fluorescence. The quenching of the tryptophyl fluorescence of model compounds and of lysozyme by iodide ion.
TL;DR: The results of the model compound study provide evidence for a mechanism that follows the classical Stern-Volmer law (1919), predominantly involving collisional quenching, and illustrate the importance of local charge and solvent viscosity.