Purification and Electrophoretic Behavior of Sugarcane Invertases

Abstract

A fair degree of sugarcane invertase purification has been achieved by techniques of differential solubility, gel filtration, and paper electrophoresis. Invertase is readily salted out with ammoniun sulfate between 38- and 52- percent saturation. Activity is largely lost during dialysis against distilled water, but is regained by passage through columns of G-200 Sephadex gel. Reactivation is attributed to removal of unknown inhibitors. Filtration did not accomplish good separation of invertase from other protein. Electrophoresis experiments showed that invertases are quite mobile compared to contaminant protein, and move quickly toward positive and negative electrodes. Two distinct invertase areas were obtained free of contaminant protein, one enzyme bearing a positive charge and the other a negative charge. Filtration and electrophoresis steps described herein achieved good enzyme purification without use of glucose or manganese, and thus avoided the appearance of reconstituted invertase.