The State of Tyrosine and Phenylalanine Residues in Proteins Analyzed by Fourth-Derivative Spectrophotometry. Histone H1 and Ribonuclease A

Abstract

The analysis of the absorption spectra of model compounds of tyrosine and phenylalanine residues by means of fourth-derivative spectrophotometry is able to separate the contribution of the 2 chromophores, thus allowing the study of each one. Fourth-derivative analysis resolves the 2 main vibrational bands of tyrosine, giving rise to 2 peaks which are sensitive to changes in the environment of the phenolic ring. The parameters obtained from the fourth-derivative spectra depended on the strength of the H-bonds formed by the OH group of tyrosine, as well as on the heterogeneity of tyrosine environments. The fourth derivative tyrosine peaks are not perturbed by broad bands, such as that arising from ionized tyrosine chromophores. The peaks arising from the phenylalanine model, although less sensitive than those of tyrosine, depended on the polarity of the environment. As a check of the method, it is applied to the study of tyrosine and phenylalanine residues of calf thymus histone H1 and bovine pancreatic RNase A.