The inactivation of α -chymotrypsin by ionizing radiations

Abstract

The inactivation of α -chymotrypsin by low-voltage X-rays and by 15 meV electrons has been studied over a range of concentrations extending from the solid enzyme to dilute solutions and the sensitivity D 37 / c , where D 37 is the dose required to cause in activation to 37 % of the original at concentration c , determined under varied circumstances. The sensitivity is constant in air over a wide range of concentrations, but in the solid state is greater by a factor of about 7. That the enhanced sensitivity in the solid state is connected with disorganization of the secondary structure is shown by the fact that after partial inactivation by irradiation the enzyme is more sensitive to in activation by heating. This view is also supported by the finding that oxygen has no significant effect on the irradiation (with 15 meV electrons) in the solid state, since there is no reason to expect that oxygen will influence the breakage of hydrogen bonds within the molecule. The sensitivities of the protease and esterase activities of the enzyme are the same, showing that only one kind of active centre is involved. The sensitivity also decreases at low concentrations of the enzyme. That this is not due to recombination of the radicals is shown by the finding that the effect is uninfluenced by varying the dose rate of the electron beam over a very wide range. An enhanced sensitivity is, however, observed in solutions from which the oxygen has been removed. It follows that secondary radicals, principally O 2 H , formed in the presence of oxygen are less effective than the primary radicals. Kinetic equations are deduced which represent the main features of this behaviour. A possible reason for the greater effectiveness of H than O 2 H is the ability of the former to penetrate into the protein molecule. It was also found that in dilute solutions containing oxygen the electron beam is more effective than the X-rays. This could be accounted for if equilibrium between the primary radicals and oxygen is not reached at the very high dose rates at which the electron pulses are delivered. In vacuo no differences in sensitiveness to the X-rays and the electrons were observed.