Phenylalanine ultraviolet absorption

Amino acids do not give any very useful ultraviolet absorption spectra unless they possess aromatic groups as in phenylalanine, tryptophan, and tyrosine. The absorption characteristics of these groups are more useful in monitoring chemical and conformational changes in proteins than they are in the simple amino acids. 

Ultraviolet absorption spectra of tryptophan (Trp), tyrosine (Tyr), and phenylalanine (Phe) at pH 6. The molar absorptivity is reflected in the extinction coefficient, with the concentration of the absorbing species expressed in moles per liter. (Source From D. B. Wetlaufer, Adv. Protein Chem. 17 303-390, 1962.)... 

Ultraviolet Absorption The ultraviolet absorption characteristics of the aromatic side chains of the primary amino acid sequence are often used to estimate an extinction coefficient for a protein. The extinction coefficient is calculated based upon the number of tyrosine, tryptophan, and phenylalanine (aromatic) residues in the protein. Phenylalanine has a relatively weaker absorptivity and may not be included in the calculations. One advantage of this approach is the fact that it is relatively nondisruptive, leaving protein conformation undisturbed. In addition, the same solution used for the concentration assessment can be removed from the cuvette and used for additional experiments. The measurement is based upon Beer s law ... 

The application of low-temperature techniques to the investigation of protein spectra in the ultraviolet region was initiated by Lavin and Northrop (1935) who investigated the ultraviolet absorption spectra of pepsin, serum albumin, and ovalbumin in glycerol, and showed that the fine structure of the protein spectrum was enhanced at — 100°C. Preliminary reports of similar work have been published by Randall and Brown (1949) on thin films of sublimed tryptophan and phenylalanine at 90°C., and by Sinsheimer et al. (1949) for tryptophan at 77.6°K. Loof-bourow and his coworkers (Sinsheimer et al., 1950) have begun publication of a series of papers reporting much more comprehensive work on the influence of low temperature on the spectra of amino acids and proteins in thin films and in solid solution. Beaven et al. (1950) have reported a few results on thin Aims of the aromatic amino acids. 

Fig. 2. Ultraviolet absorption spectra of tryptophan, tyrosine, and phenylalanine at pH 6 (from Wetlaufer >).

Fig. 6. Ultraviolet absorption spectra of phenylalanine and tryptophan (a) and tyrosine (pH 7 and pH10)(b)...

The ultraviolet absorption spectra show an extinction coefficient, which rapidly increases for wavelengths lower than 2600 A. The three licheniformins, primarily B, and less so for C, show, at about 2500 A, absorption bands corresponding to those given by phenylalanine. 

Side chains of the three aromatic amino acids phenylalanine, tyrosine, and tryptophan absorb ultraviolet light in the 240- to 300-nm region, while histidine and cystine absorb to a lesser extent. Figure 3-13 shows the absorption spectrum of a "reference compound" for tyrosine. There are three major absorption bands, the first one at 275 nm being a contributor to the well-... 

The aromatic amino acids phenylalanine, tyrosine, and tryptophan all possess absorption maxima in the near-ultraviolet (fig. 3.7). These absorption bands arise from the interaction of radiation with electrons in the aromatic rings. The near-... 

Phenylalanine s phenyl side chain classifies it as an aromatic amino acid. The aromatic amino acids, like most compounds carrying conjugated rings, exhibit strong absorption of light in the near-ultraviolet region of the spectrum (Figure 5.6). This absorption is frequently used for the analytical detection of proteins. 

The main features of the near- and far-ultraviolet spectra of the proteins are related to the absorption properties of the aromatic amino acids phenylalanine (Phe), tyrosine (Tyr), tryptophan (Trp), and histidine (His) [113,114]. The peaks observed in the absorption spectra up to 185 nm (6.70 eV) can be assigned to the excited states of the chromo-phore-acting molecules benzene, phenol, indole, and imidazole, respectively. In the present section we focus on the theoretical description of the most representative valence singlet excited states of the aromatic amino acid chromophores. As the results for benzene and phenol have been recently described [13, 46], only the results for indole, [(4) in Fig. 3] and imidazole [(5) in Fig. 3] are reviewed here [115,116]. The theoretical results support the assignment of four valence singlet states as... 

Of the twenty amino acids found in proteins, three may be classified as aromatic in character and account for virtually all of the protein absorption of ultraviolet light above 250 nm and for all of the observed luminescence. The molecular structures of the aromatic amino acids are shown in Fig. 1. The parent aromatic molecules of tryptophan, tyrosine and phenylalanine are, respectively, indole, phenol and benzene. The parents with methyl substituents in place of the amino acid side chains have the common names skatole, p-cresol and toluene, respectively. 

For a compound to be analyzed by spectrophotometry, it must absorb electromagnetic radiation, and this absorption should be distinguishable from that of other species in the sample. Biochemists assay protein solutions in the ultraviolet region at 280 nm, where the aromatic amino acids tyrosine, phenylalanine, and tryptophan (Table 11-1) have maximum absorbance. Common salts, buffers, and carbohydrates have little absorbance at this wavelength. In this section, we use Beer s law for a simple analysis and then discuss the measurement of nitrite in an aquarium. 

One research area that has benefited from extensive fluorescence investigations is the field of protein structure-function studies. There are three aromatic amino acids that absorb light in the ultraviolet spectral range, phenylalanine (Phe), tyrosine (lyr) and tryptophan (Trp). Because of their molar absorptivity the fluorescence of Tyr (2276 = 1405 m cm ) and Trp (e28o = 5579 cm" ) have... 

Note that it is the amino acids phenylalanine, tyrosine, and tryptophan which give proteins an absorption spectra in the ultraviolet. For proteins, this absorption maximum is commonly considered to be 280 nm. 

Ley and v. Englehardt (1910), Kowalski (1911), and Marsh (1924) have investigated the ultraviolet fluorescence of phenol and phenolic derivatives. They found broad emission bands extending from 2700 A to 4000 A with maxima usually in the region of 3000 A. Similar bands may be expected for tyrosine and phenylalanine and in the absence of internal quenching effects, also in proteins. A study of the ultraviolet fluorescence of aromatic amino acids and proteins would no doubt provide interesting details as regards the interaction of the aromatic residues in the protein molecules as well as help in the interpretation of the absorption spectra themselves. 

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