Identification of a mammalian glutaminyl cyclase converting glutaminyl into pyroglutamyl peptides.
Extracts from bovine pituitary were found to contain an activity catalyzing the conversion of glutaminyl peptides such as [Gln1]gonadotropin-releasing hormone, [Gln1, Gly4]thyrotropin-releasing hormone (H-Gln-His-Pro-Gly-OH), and H-Gln-Tyr-Ala-OH to the respective pyroglutamyl peptides. The TRH precursor fragment H-Lys-Arg-Gln-His-Pro-Gly-Lys-Arg-OH and the D-glutaminyl stereoisomer of H-Gln-Tyr-Ala-OH did not react under the same conditions. The conversion products were identified by Edman degradation, amino acid analysis, and reversed-phase HPLC. That this activity was exhibited by an enzyme, glutaminyl cyclase, was concluded from the protein character of the activity (revealed by its abolition with trypsin and heat), the Michaelis-Menten relationship between substrate concentration and conversion rate, and the substrate specificity. It was determined that glutaminyl cyclase had a molecular weight of 43,000-50,000, a pH optimum at pH 8, and Km and Vmax values in the range of 60-130 microM and 390-690 pmol/microgram per hr, respectively. Glutaminyl cyclase was not observed to require ATP and could be inhibited with 1.0 M ammonium chloride, which increased the Km and decreased the Vmax value. The subcellular distribution of glutaminyl cyclase corresponded to the one of peptidylglycine alpha-amidating monooxygenase believed to catalyze C-terminal amidations during posttranslational precursor processing. It was also observed that the formation of pyroglutamyl from glutaminyl peptides occurred nonenzymatically; however, the enzymatic reaction carried out with crude extract was found to be approximately 70 times faster than the nonenzymatic reaction enhanced by phosphate. It is speculated that glutaminyl cyclase may participate in the posttranslational processing of hormonal precursors to pyroglutamyl peptides.
Fischer
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WH
Spiess
J
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D000217:Acyltransferases / Q000378:metabolism*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D019881:Aminoacyltransferases; D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D005973:Glutamine / Q000378:metabolism*; D007700:Kinetics; D009842:Oligopeptides / Q000096:biosynthesis; D010902:Pituitary Gland / Q000201:enzymology*; D011499:Protein Processing, Post-Translational; D011761:Pyrrolidonecarboxylic Acid; D014357:Trypsin
D000596:Amino Acids; D009842:Oligopeptides; D005973:Glutamine; D000217:Acyltransferases; D019881:Aminoacyltransferases; C059280:glutaminyl-peptide cyclotransferase; D014357:Trypsin; D011761:Pyrrolidonecarboxylic Acid