Spectrophotometric assays for L-lysine alpha-oxidase and gamma-glutamylamine cyclotransferase.
A new assay for l-lysine alpha-oxidase is described. In this assay, the oxidized product generated from l-lysine is reacted with semicarbazide to form alpha-keto-epsilon-aminocaproate semicarbazone. Formation of the alpha-keto acid semicarbazone is continuously monitored spectrophotometrically at 248 nm (epsilon 10,160 +/- 240 M(-1) cm(-1)). The method was adapted to provide a new assay for gamma-glutamylamine cyclotransferase. This enzyme catalyzes the conversion of many l-gamma-glutamylamines to 5-oxo-l-proline and free amine. A biologically important substrate is N(epsilon)-(gamma-l-glutamyl)-l-lysine, which is converted to 5-oxo-l-proline and l-lysine by the action of gamma-glutamylamine cyclotransferase. The l-lysine generated from N(epsilon)-(gamma-l-glutamyl)-l-lysine in an endpoint assay is converted to alpha-keto epsilon-aminocaproate semicarbazone in the presence of semicarbazide, excess l-lysine alpha-oxidase, and catalase. The methods were applied to the determination of gamma-glutamylamine cyclotransferase activity of partially purified preparations of the bovine kidney enzyme and to detect gamma-glutamylamine cyclotransferase activity in rat kidney and liver homogenates.
Danson
Jedidah W
JW
Department of Neurology and Neuroscience, Weill Medical College of Cornell University, New York, New York, 10021, USA.
Trawick
Mary Lynn
ML
Cooper
Arthur J L
AJ
D000594:Amino Acid Oxidoreductases / Q000032:analysis* / Q000037:antagonists & inhibitors; D000614:Aminocaproates / Q000737:chemistry; D000818:Animals; D002227:Carbazoles / Q000737:chemistry; D002273:Carcinogens / Q000494:pharmacology; D002417:Cattle; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008239:Lysine / Q000378:metabolism; D008297:Male; D019636:Neurodegenerative Diseases / Q000201:enzymology; D051381:Rats; D011916:Rats, Inbred F344; D012663:Semicarbazides / Q000494:pharmacology; D013053:Spectrophotometry / Q000379:methods*; D005722:gamma-Glutamylcyclotransferase / Q000032:analysis*; D005723:gamma-Glutamyltransferase / Q000378:metabolism
D000614:Aminocaproates; D002227:Carbazoles; D002273:Carcinogens; D012663:Semicarbazides; C455439:alpha-keto-epsilon-aminocaproate semicarbazone; C010059:carbamylhydrazine; D000594:Amino Acid Oxidoreductases; C041123:L-lysine oxidase; D005723:gamma-Glutamyltransferase; D005722:gamma-Glutamylcyclotransferase; D008239:Lysine