title: A spectrophotometric assay for glutaminyl-peptide cyclizing enzymes.
abstract: Current assays for the glutaminyl-peptide cyclizing enzyme, glutaminyl cyclase, have several shortcomings. In this report a rapid spectrophotometric assay for cyclization of glutaminyl-peptides to pyroglutamyl-peptides by glutaminyl cyclase is described which overcomes many of these shortcomings. This coupled assay utilizes glutamate dehydrogenase, alpha-ketoglutarate and NADH to measure the ammonia released during cyclization of the dipeptide substrate Gln-Gln. Glutaminyl cyclase from bovine pituitary, partially purified by ion exchange chromatography, exhibits a Km for this substrate of 0.6 mM and a Vmax of 9.6 nmol/min/mg protein. These values are comparable to ones previously reported using other glutaminyl-peptide substrates and either radioimmunoassay or high-performance liquid chromatography to measure glutaminyl cyclase activity.
authors: Department of Chemistry, University of Southern Mississippi, Hattiesburg 39406-5043.
authors: R C
authors: RC
authors: Bateman
mesh_terms: D000217:Acyltransferases / Q000032:analysis*; D019881:Aminoacyltransferases; D000818:Animals; D002417:Cattle; D005973:Glutamine / Q000378:metabolism; D010902:Pituitary Gland / Q000201:enzymology*; D013053:Spectrophotometry / Q000379:methods*
chemical_list: D005973:Glutamine; D000217:Acyltransferases; D019881:Aminoacyltransferases; C059280:glutaminyl-peptide cyclotransferase