A Simple Method for the Preparation of Crystalline Potato Phosphorylase and Q-Enzyme

Abstract

IN the purification of potato phosphorylase¹ by ethanol precipitation², it has been found that conditions established for a particular variety of potato, harvested at a particular time of the year, are not generally applicable to other potatoes, even though these may contain as much enzyme. This is because the solubility of the enzyme is affected by its association with other constituents, the nature and quantity of which vary with the type of potato and the season. Since, in spite of this association, the enzyme is always able to react with its substrate, it was thought that the formation of the enzyme substrate complex might be utilized to free the enzyme from these other complexing molecules. Preliminary experiments in which the precipitation of phosphorylase was carried out in the presence of glucose-1-phosphate did not, however, show any appreciable change in the solubility of the enzyme. These experiments were not pursued further after it had been found that the presence of the co-substrate, amylose, results in a strong and selective precipitation of phosphorylase at a concentration of ethanol at which the enzyme is otherwise soluble², and in at least a thousandfold purification of the enzyme with respect to protein. The phosphorylase is accompanied in the precipitate by the Q-enzyme of the potato³. The enzymes are readily extracted from the precipitate and can then be precipitated from solution with ammonium sulphate in crystalline form.