Multiple forms of glutamate decarboxylase in porcine brain

Abstract

Three forms of glutamate decarboxylase from hog brain (termed alpha-, beta-, and gamma-GAD) were separated by hydrophobic interaction chromatography on phenyl-Sepharose, by isoelectric focusing, and by polyacrylamide gel electrophoresis. When rechromatographed on phenyl-Sepharose, each form migrated as a single entity, indicating that the forms are not readily interconvertible. The three forms are not different-sized aggregates of one form, since all three have the same approximate molecular weight (100,000) as determined by Sephadex G-200 chromatography. The pIs of the three forms separated by phenyl-Sepharose were determined by isoelectric focusing. The values obtained (5.3, 5.5, and 5.8 for alpha-, beta-, and gamma-GAD, respectively) were comparable to the pIs of the three peaks of activity observed upon focusing of enzyme that had been subjected to phenyl-Sepharose chromatography. These results indicate that phenyl-Sepharose chromatography and isoelectric focusing separate the same three components. When synaptosomal extracts were analyzed by phenyl-Sepharose chromatography without intervening purification steps, all three forms were present, but the proportion of beta-GAD was somewhat higher and that of gamma-GAD somewhat lower than in the usual preparations.