Hydrolytic enzymes involved in glycogen metabolism

Abstract

Acid and neutral a-glucosidases, with apparent molecular weights of 179,900 and 27,500, respectively, have been partially purified from rat liver tissue. The neutral enzyme is contaminated with an acid a-glucosidase of similar molecular weight. The high molecular weight acid a-glucosidase can be dissociated into inactive sub-units by m-urea. Liver fractionation studies confirm that the high molecular weight acid a-glucosidase is located in the lysosomal fraction, in soluble and membrane-bound forms, whereas the neutral enzyme (with its associated acid enzyme) is present in the microsomal fraction. A large-scale preparative technique lias been devised to purify rat liver lysosomes by rate-zonal centrifugation through a sucrose-density gradient. The purity of the organelles prepared by this method have been studied by biochemical analyses and electron microscopy.The action of various hormones on rat liver a-glucosidases lias been examined. Cortisone acetate was observed to activate rat liver acid a-glucosidase (but not the neutral enzyme) in vivo, liver slices and purified lysosomes and in human liver tissue. It was noted that for activation of the enzyme in lysosomes to occur, intact organelles were necessary. When C-4-cortisone acetate was incubated with purified lysosomes, 78% of the total activity recovered in the organelles was present in the membrane fraction and thus suggested that the membrane played an important role in the activation phenomenon. In addition, it was only the soluble enzyme, and not the membrane-bound enzyme, that was activated. Changes in the structure of the steroid resulted in a loss of the activation property. Some other lysosomal hydrolases were not affected by cortisone acetate. K and V determinations suggested that cortisone acetateacted by increasing the catalytic efficiency of the lysosomal acid a-glucosidase. However, the intralysosomal glycogen level in vivo was not lowered when the rats were treated with cortisone acetate. Possible mechanisms for the enzyme activation are discussed.