title: The NH2-terminal proregion of peptidylglycine alpha-amidating monooxygenase facilitates the secretion of soluble proteins.
abstract: A highly conserved ten amino acid proregion separates the peptidylglycine alpha-hydroxylating monooxygenase (PHM) domain of the bifunctional peptidylglycine alpha-amidating monooxygenase (PAM) protein from the NH2-terminal signal peptide; propeptides with amino acid sequences similar to the PAM proregion have been identified in other secreted proteins. In AtT-20 cells, but not in human embryonic kidney (hEK)-293 cells, an endogenous endoprotease acting at a site distal to the trans-Golgi network efficiently removes the propeptide from stably transfected monofunctional PHM (PHMs). We constructed a mutant PHM protein (delta ProPHMs) in which the proregion was deleted and the signal peptide joined directly to the monooxygenase domain. Newly synthesized, enzymatically active delta ProPHMs was secreted from both AtT-20 cells and hEK-293 cells more slowly than PHMs. In endocrine cells, the proregion was not required for storage in regulated secretory granules. We transferred the PAM proregion to prohormone convertase 2 (PC2), another soluble constituent of secretory granules, to determine whether the effect of the proregion were transferrable. In both AtT-20 cells and hEK-293 cells, the PAM/PC2 fusion molecule was able to exit the endoplasmic reticulum more rapidly than PC2.
authors.affiliation: Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: R E
authors.initials: RE
authors.lastname: Mains
authors.forename: S L
authors.initials: SL
authors.lastname: Milgram
authors.forename: H T
authors.initials: HT
authors.lastname: Keutmann
authors.forename: B A
authors.initials: BA
authors.lastname: Eipper
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D003594:Cytoplasmic Granules / Q000201:enzymology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004792:Enzyme Precursors / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000196:embryology; D051379:Mice; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000502:physiology*; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D010911:Pituitary Neoplasms / Q000473:pathology; D011333:Pro-Opiomelanocortin / Q000737:chemistry / Q000378:metabolism; D045706:Proprotein Convertase 2; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000737:chemistry; D011506:Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012995:Solubility; D013381:Subtilisins / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D004792:Enzyme Precursors; D009097:Multienzyme Complexes; D021382:Protein Sorting Signals; D011506:Proteins; D011993:Recombinant Fusion Proteins; D011333:Pro-Opiomelanocortin; D006899:Mixed Function Oxygenases; C035189:peptidylglycine monooxygenase; D013381:Subtilisins; D045706:Proprotein Convertase 2