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Zhengping Yi M.A. QasimSabiha Qasim T.L. WarringtonMichael Laskowski Jr. 《Biochimica et Biophysica Acta (BBA)/General Subjects》2006
We have used UV difference spectroscopy and fluorescence spectroscopy to study the perturbation by β-cyclodextrin of tyrosyl or tryptophyl residues located at each of the 10 variable consensus contact positions in the third domain of turkey ovomucoid. The goal was to monitor the accessibility of the side chain rings of these residues when located at these positions. The results indicated that the tyrosyl or tryptophyl rings are most highly exposed when located in the P1 position followed by the P4 position. It was possible to determine the association constants for β-cyclodextrin binding at these positions. When located at the P2, P5, P6 and P3′ positions, the rings of the tyrosyl or tryptophyl residues were exposed but less so than at the P1 or P4 positions. By contrast, when located at the P1′, P2′, P14′ and P18′ positions, the tyrosyl or tryptophyl residues were insufficiently exposed to be perturbed by β-cyclodextrin, although they reacted positively to dimethyl sulfoxide solvent perturbation. These findings indicate that β-cyclodextrin perturbation provides a convenient way to detect highly exposed tyrosyls or tryptophyls in proteins. Furthermore, we evaluated the ability of β-cyclodextrin to inhibit the interaction of turkey ovomucoid third domain variants with different P1 residues. The results showed that the presence of β-cyclodextrin had little effect on the association constant when the P1 residue was a glycyl residue, but greatly decreased the association constant when the P1 residue was a tyrosyl or tryptophyl residue. Thus, β-cyclodextrin may be used to selectively modulate the interaction between proteinase inhibitors and their cognate enzymes. 相似文献
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Ciro Balestrieri Giovanni Colonna Alfonso Giovane Gaetano Irace Luigi Servillo 《Analytical biochemistry》1980,106(1):49-54
Ionization of the phenolic group of N-acetyltyrosynamide has been studied using second-derivative spectroscopy. At pH 12.5 the second-derivative spectrum of the model compound revealed the presence of derivative bands in a spectral region (between 250 and 270 nm) where interference coming from other ultraviolet-absorbing chromophores is negligible. One of these peaks (260-nm peak) has been employed for the determination of tyrosyl groups in mixtures containing the aromatic amino acids. 相似文献
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The states of tyrosyl and tryptophyl residues in a protein proteinase inhibitor (Streptomyces subtilisin inhibitor 总被引:1,自引:0,他引:1
The states of tyrosyl and tryptophyl residues of a dimeric protein proteinase inhibitor, Streptomyces subtilisin inhibitor (Sato, S & Murao, S. (1973), Agric. Biol. Chem. 37, 1067) were studies by solvent perturbation difference spectroscopy with methanol, ethylene glycol, polyethylene glycol, and deuterium oxide as perturbants, and by spectrophotometric titration at alkaline pH. It appeared that all three tyrosyl residues per monomer of the inhibitor were exposed on the surface of the molecule, and their apparent pK values were estimated separately to be 9.58, 11.10, and 12.42. The single tryptophyl residue per monomer of the inhibitor appeared to be partially buried in the protein molecule. 相似文献
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G B Villanueva C W Batt E Sausville V Tortorelli K Cubelli W Brunner 《Biochemical and biophysical research communications》1975,67(1):359-365
Chemical modifications of tyrosine and tryptophan residues in diisopropylphosphoryl-thrombin (DIP-thrombin) and benzamidine-inhibited thrombin (BA-thrombin) by N-acetylimidazole and hydrogen peroxide-dioxane mixture indicate the burial of two tyrosyl and two tryptophyl residues relative to the active enzyme. During inhibition the circular dichroism spectra in the peptide-absorbing region is apparently unchanged while small detectable changes are observed in the aromatic region. It is concluded that tryptophan and tyrosine residues are part of the structural features of the active center of thrombin but they do not play active roles in the catalytic process. 相似文献
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Ultraviolet difference absorption spectra produced by ethylene glycol were measured for hen lysozyme [EC 3.2.1.17] and bovine chymotrypsinogen. N-Acetyl-L-tryptophanamide and N-acetyl-L-tyrosinamide were employed as model compounds for tryptophyl and tyrosyl residues, respectively, and their ultraviolet difference spectra were also measured as a function of ethylene glycol concentration. By comparison of the slopes of plots of molar difference extinction coefficients (delta epsilon) versus ethylene glycol concentration for the proteins with those of the model compounds at peak positions (291-293 and 284-287 nm) in the difference spectra, the average number of tyrosyl as well as tryptophyl residues in exposed states could be estimated. The results gave 2.7 tryptophyl and 1.9 tyrosyl residues exposed for lysozyme at pH 2.1 and 2.6 tryptophyl and 3.4 tyrosyl residues exposed for chymotrypsinogen at pH 5.4. The somewhat higher tyrosyl exposure of chymotrypsinogen, compared with the findings from spectrophotometric titration and chemical modification, was not unexpected, because delta epsilon285 was larger than delta epsilon292, and the situation is discussed with reference to preferential interaction of ethylene glycol with the tyrosyl residues and/or side chains in the vicinity of the chromophore in the protein. The procedure employed in the present work seems to be suitable for estimation of the average number of exposed tryptophyl and tyrosyl residues in tryptophan-rich proteins. The effects of ethylene glycol on the circular dichroism spectra of lysozyme at pH 2.1 and chymotrypsinogen at pH 5.4 were also investigated. At high ethylene glycol concentrations, both proteins were found to undergo conformational changes in the direction of more ordered structures, presumably more helical for lysozyme and more beta-structured for chymotrypsinogen. 相似文献