Inactivation of Snake Venom L-Amino Acid Oxidase by Freezing

Abstract

Abstract The l-amino acid oxidase of Crotalus adamanteus is inactivated by storage in the frozen state at temperatures between -5° and -60°, with maximal inactivation being observed at approximately -20°. The rate of inactivation is dependent on the pH of storage and on the ionic composition of the medium. The inactivation is accompanied by changes in the visible absorption spectrum and by a decrease in the rate of photoreduction with ethylenediaminetetraacetate. The frozen, inactivated enzyme in most cases (depending on the conditions of storage) can be reactivated completely by heating at pH 5, with a full return of the spectrum of native enzyme. The reactivation process is extremely dependent on temperature, with a calculated ΔH of approximately 40,000 cal per mole. The results are consistent with the hypothesis that the inactivation is due to a limited conformational change of the enzyme structure, particularly in the vicinity of the flavin prosthetic group. The similarities and differences between this type of inactivation and the reversible heat inactivation of this enzyme observed by previous workers are discussed.