Human serum albumin tryptophan

Soltes, L., Sebille, B. (1997). Reversible binding interactions between the tryptophan enantiomers and albumins of different animal species as determined by novel high performance liquid chromatographic methods an attempt to localize the d- and L-tryptophan binding sites on the human serum albumin polypeptide chain by using protein fragments. Chirality 9, 373-379. 

Derrick studied the interaction of L-tryptophan and ibuprofen with human serum albumin (HSA),74 which is an abundant transport blood protein capable of binding efficiently several species.75 They acquired 1H NMR spectra of L-Tryptophan-HSA system for different ligand protein molar ratios, that is 3 1, 5 1, 7 1 and 10 1. The aromatic resonances of L-Tryptophan are difficult to be observed due to the overlap with HSA signals, even at 10 1 molar ratio, so that the spectral subtraction was performed. D values of L-Tryptophan were calculated by integration of the subtracted spectra and were in good agreement with those predicted by computer simulations. In the case of ibuprofen, only for 140 1 molar ratio, the resonances of ibuprofen are clearly visible also in this case, the... 

Investigations of the effects of UV- and hypochlorite-induced oxidative modification of 20 amino acids and human serum albumin (HSA) on their antiradical properties showed unexpected results [36], Seven amino acids (cystine, histidine, methionine, phenylalanine, serine, tryptophan, and tyrosine) and HSA developed ACW following oxidation (see examples in Fig. 14). The fresh (produced in 1998) HSA from Serva had no antiradical capacity, but it acquired this quality during irradiation. The out-of-date HSA sample (Dessau, GDR, 1987, expiration date 7/1/1992) showed a remarkable ACW even in an unirradiated state. 

Figure 2.11. The dependence of the position of the fluorescence spectrum maximum on excitation wavelength for tryptophan in a model medium (glycerol) at different temperatures (a) and singletryptophan proteins (b). 1, Whiting parvalbumin, pH 6.S in the presence of Ca2+ ions 2, ribonuclease Th pH 6.5 3, ribonuclease C2, pH 6.5 4, human serum albumin, pH 7.0, +10"4 M sodium dodecyl sulfate 5, human serum albumin, pH 3.2 6, melittin, pH 7.5, +0.15 M NaCl 7, protease inhibitor IT-AJ from Actinomyces janthinus, pH 2.9 8, human serum albumin, pH 7.0 9, -casein, pH 7.5 10, protease inhibitor IT-AJ, pH 7.0 11, basic myelin protein, pH 7.0 12, melittin in water. The dashed line is the absorption spectrum of tryptophan.

At 77 K the position of the 0-0 band is generally blue shifted for exposed tryptophans and red shifted for buried tryptophans. Along with a shift in wavelength to the red, the phosphorescence lifetime decreases.(28) The single tryptophan of human serum albumin shows red-shifted phosphorescence and D - L triplet zero-field splitting, indicating that it is in a hydrophobic environment. 29 ... 

K. L. Bell and H. C. Brenner, Phosphorescence and optically detected magnetic resonance study of the tryptophan residue in human serum albumin, Biochemistry 21, 799-804 (1982). 

Yang, J. and Hage, D.S., Effect of mobile phase composition on the binding kinetics of chiral solutes on a protein-based HPLC column Interactions of d- and L-tryptophan with immobilized human serum albumin, J. Chromatogr. A, 766, 15-25, 1997. 

In this section, we review our first examinations of tryptophan probing sensitivity and water dynamics in a series of important model systems from simple to complex, which range from a tripeptide [70], to a prototype membrane protein melittin [70], to a common drug transporter human serum albumin [71], and to lipid interface of a nanochannel [86]. At the end, we also give a special case that using indole moiety of tryptophan probes supramolecule crown ether solvation, and we observed solvent-induced supramolecule folding [87]. The obtained solvation dynamics in these systems are linked to properties or functions of these biological-relevant macromolecules. 

Peroxynitrite is a nonspecific oxidant that reacts with all classes of biomolecules depleting low-molecular-weight antioxidants, initiating lipid peroxidation, damaging nucleic acids and proteins. Its reactions are much slower than those of the hydroxyl radical but are faster than those of hydrogen peroxide. Comparison of peroxynitrite reactivity with various amino acid residues of human serum albumin have shown that cysteine, methionine, and tryptophan are the most reactive... 

Figure 13 shows these changes in extinction for the NBS titration of tryptophan in bovine serum albumin (Ramachandran and Witkop, 1959), which is dissolved in 10.0 M urea solution in order to make the tryptophan units accessible. Another convenient way of picturing the changes in extinction is shown in Fig. 14 (Peters, 1959). Here one recognizes at a glance that ribonuclease contains no tryptophan. Based on a value of 2.8 X 10 for the amplitude in drop of molar absorption of free tryptophan it was concluded that human serum albumin (HSA) contains one, bovine serum albumin (BSA) two, and ovalbumin probably four rather than three tryptophan units.

B. Subfile and N. Thuaud, Determination of tryptophan-human serum albumin binding from retention data and separation of tryptophan enantiomers by high-performance liquid chromatography, J. Litj. Chromatogr., 3 299 (1980). 

I. Fitos and M. Simonyi, Investigation of binding of tryptophan enantiomers to human serum albumin, Acta Biochim. Biophys. Hung., 21 237 (1986). 

W. E. Muller and U. WoUert, Benzodiazepines, specific competitors for the binding of L-tryptophan to human serum albumin, Naunyn-Schmiedeberg s Arch. Pharmacd., 288 17 (1975). 

For human serum albumin Tanford (1950) found by spectrophotometry that the ionization of the tyrosine hydroxyl groups was completely reversible up to pH 12. Measurements at the wavelength of the tyrosine anion maximum (2930 A.), uncorrected for the small tryptophan contribution, gave a pK of 11.7 for this process. Both the ultraviolet absorption and titration data for this protein could be quantitatively interpreted on the basis of complete freedom of all the 18 tyrosine hydroxyl groups in the molecule to ionize. In this respect human serum albumin thus resembles insulin and not ovalbumin. 

The photophysical parameters such as (a) true absorption cross section (at 266 nm) of tryptophan and intersystem crossing rate in single-tiyptophan-containing protein human serum albumin and (b) true absorption cross section (at 266 nm) of tryptophans and rate of energy transfer between them in two-tryptophan-containing protein bovine serum albumin have been determined. 

In this work, the solutions of human serum albumin (HSA) (>96%, Sigma) and of bovine serum albumin (BSA) (>98%, MP Biomedicals) in a phosphate buffer (0.01 M, pH 7.4) have been used. The proteins concentrations were lO- (absorption spectra measurement) and 10- M (fluorescence measurement at the nanosecond laser fluorimeter). All of the experiments were performed at a temperature of 25 1 °C. The structure and biological functions of HSA and BSA can be found in (Peters, 1996). Tryptophan, tyrosine, and phenylalanine (with relative contents of 1 18 31 in HSA and 2 20 27 in BSA) are the absorption groups in these proteins (as in many other natural proteins). The tyrosine fluorescence in HSA and BSA (as in many other natural proteins) is quenched due to the effect of adjacent peptide bonds, polar groups (such as CO, NH2), and other factors, and phenylalanine has a low fluorescence quantum yield (0.03) (Permyakov, 1992). Therefore, the fluorescence signal in these proteins is determined mainly by tryptophan groups. In that case the fluorescence, registered in nonlinear and kinetic laser fluorimetry measurements, correspond to tryptophan residues (this fact will be used in Section 6.1). 

Let us discuss the main results of this Section. First of all, it is interesting to compare the true value of the absorption cross section obtained for tryptophan in the FP mRFPl with the values for tryptophan in an aqueous solution (1.6xl0 i7 cm (Banishev et al., 2008)), human serum albumin (1.3xl0 i7 cm ), and bovine serum albumin (ffD=lxlO i7 cm and (Ta=3x10"17 cm ), which were determined in previous Section. One can see that the values for tryptophans in proteins are different and do not equal to the value for a free tryptophan, as is often assumed. 1 want to emphasize that the lifetime of the excited state of the donor (tryptophan) Td in the absence of the acceptor (chromophore) has been obtained without the removal of the acceptor (as it is often supposed when determine the energy transfer... 

Human interferon-o2a, interleukin-2 receptor, human IgG, human serum albumin, dehydrogenases glucose-6-phosphate dehydrogenase, foanate dehydrogenase, pyruvate decarboxylase, S-oxynitrilase, bovine liver catalase, recombinant protein G, plasmid DMA, human blood coagulation factor VIII, enantiomers of kynurenine, tryptophan, beta-blockers, practolol, thic ntal... 

Calculation of a Distance from FRET Data The protein human serum albumin (HSA) has a single tryptophan residue at position 214. HSA was labeled with an anthra-niloyl group placed covalently on cysteine-34. Bnission... 

Figure 5 An example of a zonal elution experiment, in which small injections of L-tryptophan are made on to an immobilized human serum albumin column in the presence of increasing amounts of phenytoin in the mobile phase.

Fig. 3 Typical plots of (a) total plate height (//tot) and (b) the plate height contribution due to stationary phase mass transfer (//s) for injections of D-tryptophan at various flow rates onto an immobilized human serum albumin column. Symbols u, linear velocity kf, retention factor.

Chattopadhyay, A. Tian, T. Kortum, L. Hage, D.S. Development of tryptophan-modified human serum albumin columns for site-specific studies of dmg-protein interactions hy high-performance affinity chromatography. J. Chromatogr. B, 1998, 715, 183 190. 

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