title: Identification, purification, and characterization of the molecular forms of Aplysia californica peptidylglycine alpha-amidating enzyme.
abstract: Peptidylglycine alpha-amidating enzyme (PAM; EC 1.14.17.3) is responsible for the conversion of peptides with a COOH-terminal glycine into alpha-amidated peptides, a posttranslational modification often required for biological activity and/or increased stability. Such an activity able to convert the model peptide D-Tyr-Val-Gly into D-Tyr-Val-amide was found to be present in the marine mollusk Aplysia californica. Examination of this amidating activity as well as its immunoreactivity demonstrates that (1) it can be found mainly in the atrial gland, heart, and CNS but is barely detectable in the hepatopancreas and gonads, (2) it requires as essential cofactors copper, molecular oxygen, and ascorbate, and (3) it exists in at least two molecular forms, a soluble and a membrane-bound form. Purification of this activity from the atrial gland was accomplished using Cu(2+)-chelating Sepharose, gel permeation, and hydroxyapatite chromatography. In addition, using polyclonal antibodies raised against various parts of the rat amidating enzyme, we demonstrate that numerous immunologically recognized regions are conserved in both the soluble and membrane-bound Aplysia californica PAM.
authors.affiliation: Laboratory of Neuropeptides Structure and Metabolism, Clinical Research Institute of Montréal, Québec, Canada.
authors.forename: A
authors.initials: A
authors.lastname: Boudreault
authors.forename: V F
authors.initials: VF
authors.lastname: Castellucci
authors.forename: M
authors.initials: M
authors.lastname: Chrétien
authors.forename: C
authors.initials: C
authors.lastname: Lazure
mesh_terms: D000818:Animals; D001048:Aplysia / Q000201:enzymology*; D001345:Autoradiography; D015153:Blotting, Western; D003300:Copper / Q000737:chemistry; D004702:Endocrine Glands / Q000737:chemistry / Q000201:enzymology; D008322:Mammals; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D009097:Multienzyme Complexes; D051381:Rats; D012685:Sepharose; D013552:Swine
chemical_list: D009097:Multienzyme Complexes; D003300:Copper; D012685:Sepharose; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase