52] Glutaminase, asparaginase, and α-keto acid-ω-amidase

Abstract

Publisher Summary This chapter discusses the determination of L-glutaminase from Escherichia coli , L-asparaginase from guinea pig serum, and α -keto acid- ω -amidase from rat liver. In the case of the determination of L-glutaminase from Escherichia coli , enzyme activity may be measured by determinations of ammonia or of L-glutamate. The assay procedure described is based on determination of the rate of ammonia liberation. Enzyme solution (0.25 ml.) and L-glutamine (0.25 ml.) are incubated for 5 to 60 minutes at 37°. The reaction is stopped by addition of 0.5 ml. of 15% trichloroacetic acid. Ammonia is determined by aeration in the presence of potassium carbonate into sulfuric acid traps. Blanks containing glutamine alone and enzyme preparation alone are employed. In the case of determination of L-asparaginase from guinea pig serum, enzyme activity may be conveniently measured by determinations of ammonia. Enzyme solution (0.5 ml.), substrate (0.5 ml.), and buffer (1.0 ml.) are mixed and incubated for 5 to 60 minutes at 37 °. The reaction is stopped by addition of 0.5 ml. of 15% trichloroacetic acid, and ammonia is determined. In the case of determination of α -keto acid- ω -amidase from rat liver, the reaction is followed by determination of the ammonia or α -keto acid formed.