title: Naturally occurring active N-domain of human angiotensin I-converting enzyme.
abstract: Angiotensin I-converting enzyme (ACE, kininase II) is a single-chain protein containing two active site domains (named N- and C-domains according to position in the chain). ACE is bound to plasma membranes by its C-terminal hydrophobic transmembrane anchor. Ileal fluid, rich in ACE activity, obtained from patients after surgical colectomy was used as the source. Column chromatography, including modified affinity chromatography on lisinopril-Sepharose, yielded homogeneous ACE after only a 45-fold purification. N-terminal sequencing of ileal ACE and partial sequencing of CNBr fragments revealed the presence of an intact N terminus but only a single N-domain active site, ending between residues 443 and 559. Thus, ileal-fluid ACE is a unique enzyme differing from the widely distributed two-domain somatic enzyme or the single C-domain testicular (germinal) ACE. The molecular mass of ileal ACE is 108 kDa and when deglycosylated, the molecular mass is 68 kDa, indicating extensive glycosylation (37% by weight). In agreement with the results reported with recombinant variants of ACE, the ileal enzyme is less Cl(-)-dependent than somatic ACE; release of the C-terminal dipeptide from a peptide substrate was optimal in only 10 mM Cl-. In addition to hydrolyzing at the C-terminal end of peptides, ileal ACE efficiently cleaved the protected N-terminal tripeptide from the luteinizing hormone-releasing hormone and its congener 6-31 times faster, depending on the Cl- concentration, than the C-domain in recombinant testicular ACE. Thus we have isolated an active human ACE consisting of a single N-domain. We suggest that there is a bridge section of about 100 amino acids between the active N- and C-domains of somatic ACE where it may be proteolytically cleaved to liberate the active N-domain. These findings have potential relevance and importance in the therapeutic application of ACE inhibitors.
authors.affiliation: Department of Pharmacology, University of Illinois College of Medicine, Chicago 60612.
authors.forename: P A
authors.initials: PA
authors.lastname: Deddish
authors.forename: J
authors.initials: J
authors.lastname: Wang
authors.forename: B
authors.initials: B
authors.lastname: Michel
authors.forename: P W
authors.initials: PW
authors.lastname: Morris
authors.forename: N O
authors.initials: NO
authors.lastname: Davidson
authors.forename: R A
authors.initials: RA
authors.lastname: Skidgel
authors.forename: E G
authors.initials: EG
authors.lastname: Erdös
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003082:Colectomy; D003488:Cyanogen Bromide; D007987:Gonadotropin-Releasing Hormone / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D007082:Ileum / Q000201:enzymology*; D007419:Intestinal Secretions / Q000201:enzymology*; D007668:Kidney / Q000201:enzymology; D008297:Male; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D007703:Peptidyl-Dipeptidase A / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D011817:Rabbits; D017421:Sequence Analysis; D013379:Substrate Specificity; D013737:Testis / Q000201:enzymology; D017329:Triptorelin Pamoate / Q000031:analogs & derivatives*
chemical_list: D009842:Oligopeptides; D010446:Peptide Fragments; D017329:Triptorelin Pamoate; C010980:hippuryl-histidyl-leucine; D007987:Gonadotropin-Releasing Hormone; C019176:Tryptal; D007703:Peptidyl-Dipeptidase A; D003488:Cyanogen Bromide