Effect of temperature on X-ray-induced light emission from powders of amino acids and trypsin

The following was observed in studies of the effect of temperature on the intensity of X-ray-induced emission from powders of aromatic amino acids and proteins:. The thermal coefficients for quenching of both fluorescence and phosphorescence are much different at temperatures below about 150 °K and above 240 °K. The ratios of the coefficients measured in the two temperature regions varied from 3 to 50 in the four compounds studied. The E_q for tryptophan fluorescence in powders exposed to X-rays near 300 °K is much less than the coefficient for quenching ultraviolet-induced fluorescence from tryptophan in solution. The quenching coefficient for fluorescence is invariably smaller than that for phosphorescence. The E_q = 0.21 ± .07 eV per molecule for the quenching of trypsin phosphorescence in the range T > 200 °K is essentially the same as that for tyrosine phosphorescence. This value is approximately twice that of 0.1 eV per molecule for the thermal enhancement of trypsin inactivation by ionizing radiations in the same temperature range. Crude measurements suggest that the thermal coefficient for quenching of trypsin fluorescence is also less than the corresponding activation energies for trypsin inactivation. Analysis of the data has led to the following conclusions:. (1) Quenching is a function of the macromolecular organization of proteins. (2) Thermal enhancement of enzyme inactivation yields does not depend critically upon indiscriminate quenching to prevent emissive loss of potential inactivating energy from the irradiated molecules. (3) Either the interaction of specific excited aromatic residues with radiosensitive structures does not provide a large contribution to enzyme inactivation or the specific interactions account for only a small fraction of total quenching. The present results emphasize again the differences between the excitation events produced by X-ray-induced electrons and low-energy ultraviolet. Although trypsin fluorescence appears to be from tryptophan, X-rays cause a great enhancement in trypsin of phosphorescence typical of that from tyrosine: excitations by "ionizing" radiations must be involved in intersystem crossing with high probability. "Red" components not found in ultraviolet-induced emission are also observed.