An L-Aspartate: 2-Oxoglutarate Aminotransferase Reference Material from Human Erythrocytes: Preparation and Characterization

Abstract

L-Aspartate: 2-oxoglutarate aminotransferase (EC 2.6.1.1) from human erythrocytes is purified for use as a reference material in clinical assays for the enzyme. The procedure involves heat treatment in the presence of a substrate (2-oxoglutarate), ammonium sulfate precipitation, and fractionation with n-butanol. The material is further purified by anion- and cation-exchange chromatography. Specific activity obtainable by a typical preparation is 110 µmol min^-1 mg^-1 at 30°C. Identical behavior is demonstrated between enzyme in human serum and purified erythrocyte enzyme when substrate concentrations, temperature, and pH are varied. A lyophilized preparation at 4°C is stable for longer than 400 days. Data regarding purity and behavior in currently used clinical assays is described.