What causes the variation of polarization degree across the emission spectrum of proteins?

A gradual decrease in fluorescence polarization across the emission spectrum on increase in wavelength has been recorded for a number of proteins and also for tryptophan, N-acetyltryptophan and glycyltryptophan. Various factors responsible for this dependence have been analyzed. It is shown that if the emission originates from both the 1La and 1Lb states, the position and form of the fluorescence spectrum polarization components as well as the slope of the dependence of the degree of polarization upon emission wavelength must always vary with the excitation wavelength. However, this condition, although necessary, is not enough to prove the participation of 1Lb in emission. The dependence of the form of the emission polarization spectrum upon excitation wavelength obtained for some proteins is explained by tyrosine residues contributing to the emission. Consequently, there are no reasons for assuming that the 1Lb oscillator participates in emission. It has been observed that for individual emitting centres, the slope of the dependence of the degree of polarization upon emission wavelength is determined by alteration of the vibrational substates, between which the transition with radiation takes place. The heterogeneity in the microenvironment properties of separate tryptophan residues in multitryptophan proteins and the existence, under certain conditions, of a correlation between the radiative lifetime of the emitting centre (determining the degree of the emission polarization) and the completeness of the microenvironment orientational relaxation (determining the emitted quantum of energy) can also affect the slope of this dependence.