Tyrosine fine structure

Further studies on proteins have been carried out by Haas et al. (1951a) using low-temperature techniques to improve the resolution of the fine-structure. These methods made it possible to recognize the absorption maximum of dopa (3 4-dihydroxyphenylalanine) at 2820 A. which appears in the spectrum as a progressive enhancement of the tyrosine fine-structure maximum at the same wavelength dopa is apparently a characteristic intermediate in the oxidation of combined tyrosine or tyrosine derivatives, though not of free tyrosine. 

The two transitions for tryptophan exhibit distinct features, which lead to quite different CD bands. The La transition is broad, relatively featureless, and intense (Fig. B3.5.2). The Lb transition is weaker but exhibits fine structure (Fig. B3.5.2) similar to that of tyrosine. It can be masked by or superimposed on the La transition. Interactions similar to those listed for tyrosine strongly affect the peak wavelengths and intensities. The presence of bands above 285 nm is diagnostic for tryptophan in a specific environment interactions that cause the Lb transition to be shifted to the red (or high-wavelength) end of the spectrum result in bands as high as 310 nm which are highly conformation-specific. The two transitions may be affected quite independently by interactions, and their combination can thereby result in four distinct types of spectrum (Strickland, 1974). 

Disulfide bonds may exhibit a broad band of ellipticity in the range of 240 to 350 nm. This can be confused with the band associated with the La transition of tryptophan but, in cases where a disulfide makes a significant contribution to the CD, it can be recognized by its ellipticity above 320 nm. It can augment or diminish the apparent intensity of tryptophan and tyrosine contributions without being recognized as such and without fine structure necessarily being lost. 

Because the 2570 A band of phenylalanine is weak, it is often obscured in proteins by the much stronger tyrosine and tryptophan absorptions. It is occasionally visualized in protein spectra as ripples (fine structure) in the spectral region 2500-2700 A. These ripples can be amplified by the difference spectral technique, as is shown in Fig. 13. A typical phenylalanine difference spectrum, obtained in a comparison of the isoelectric amino acid with a solution of the same concentration at pH 1 is shown in Fig. 12. Difference spectra for phenylalanine in various solvents have been measured by Bigelow and Geschwind (1960), Yanari and Bovey (1960), and Donovan et al. (1961). Fluorescence activation and emission spectra for phenylalanine were measured by Teale and Weber (1957). 

It may be readily detected spectrographically in proteins, even in presence of tyrosine and tryptophan, by means such as the moving-plate method (Holiday, 1937, 1950a). A source giving a continuous spectrum is essential to show up the fine structure bands which reveal its presence. 

In studies of the recombination of heme with globin Jope,. lope and O Brien (1949) have used the concept of bound tyrosine hydroxyl groups as a criterion of native character in globin, based on the spectropho-tometric study of the alkaline ionization process. They were able to show that globin preparations which were native according to this criterion could be classified as denatured on the basis of the position of their tryptophan fine-structure bands when recombined with heme. These results led them to suggest that the process of denaturation could be separated into several stages, even by spectroscopic techniques alone (see also Jope, 1949). 

Inspection of the absorption curves of proteins in alkali shows that the estimate of shift should be possible. In Fig. 11 are shown for comparison curves of various mixtures of tyrosine and tryptophan in V/10 alkali together with curves of some proteins under the same conditions. It can be seen at once that a fair estimate of the tyrosine/tryptophan ratio may be obtained by inspection of the head of the band. By the moving-plate method the position of the tryptophan fine-structure band, which forms one maximum of the curve can be estimated to +1 A. By standard photoelectric spectrophotometry it can be estimated to +2-5 A. 

Many biologically active polypeptides have been shown to possess CD bands in the wavelength region above 270 nm. In peptide hormones and antibiotics and in non-conjugate proteins these bands are due to tryptophan and tyrosine residues, the only chromophores which absorb in this spectral region. Because of the weakness and spectral overlap of these Cotton effects, their structure is usually imperfectly resolved. However, in recent years some progress in analyzing the fine structure of the CD spectra of tryptophan and derivatives has been reported (7 9). 

X0vanadates, which may result in irreversible inhibition of some protein tyrosine phosphatases, as opposed to the readily reversible phosphatase inhibition seen with vanadates. In addition, vanadate s substitution for phosphate within the enzyme structure may result in formation of a transition state analog of protein tyrosine phosphatase, thus changing the kinetics of tyrosine kinase activation, and effectively acting as a means of fine tuning the intracellular balance between phosphatase inhibition and kinase activation. ... 

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