title: Cloning and expression of an evolutionary conserved single-domain angiotensin converting enzyme from Drosophila melanogaster.
abstract: Mammalian somatic angiotensin converting enzyme (EC 3.4.15.1, ACE) consists of two highly homologous (N- and C-) domains encoded by a duplicated gene. We have identified an apparent single-domain (67 kDa) insect angiotensin converting enzyme (AnCE) in embryos of Drosophila melanogaster which converts angiotensin I to angiotensin II (Km, 365 microM), removes Phe-Arg from the C terminus of bradykinin (Km, 22 microM), and is inhibited by ACE inhibitors, captopril (IC50 = 1.1 x 10(-9) M) and trandolaprilat (IC50 = 1.6 x 10(-8) M). We also report the cloning and expression of a Drosophila AnCE cDNA which codes for a single-domain 615-amino acid protein with a predicted 17-amino acid signal peptide and regions with high levels of homology to both the N- and C-domains of mammalian somatic ACE, especially around the active site consensus sequence. Northern analysis identified a single 2.1-kilobase mRNA in Drosophila embryos, and Southern analysis of Drosophila genomic DNA indicates that the insect gene is not duplicated. When expressed in COS-7 cells, the AnCE protein is a secreted enzyme, which converts angiotensin I to angiotensin II and is inhibited by captopril (IC50 = 5.6 x 10(-9) M) and trandolaprilat (IC50 = 2 x 10(-8) M). The evolutionary significance of these results is discussed.
authors.affiliation: Department of Pure & Applied Biology, University of Leeds, United Kingdom.
authors.forename: M J
authors.initials: MJ
authors.lastname: Cornell
authors.forename: T A
authors.initials: TA
authors.lastname: Williams
authors.forename: N S
authors.initials: NS
authors.lastname: Lamango
authors.forename: D
authors.initials: D
authors.lastname: Coates
authors.forename: P
authors.initials: P
authors.lastname: Corvol
authors.forename: F
authors.initials: F
authors.lastname: Soubrier
authors.forename: J
authors.initials: J
authors.lastname: Hoheisel
authors.forename: H
authors.initials: H
authors.lastname: Lehrach
authors.forename: R E
authors.initials: RE
authors.lastname: Isaac
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005075:Biological Evolution; D002478:Cells, Cultured; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004331:Drosophila melanogaster / Q000201:enzymology*; D008969:Molecular Sequence Data; D007703:Peptidyl-Dipeptidase A / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D011994:Recombinant Proteins; D007703:Peptidyl-Dipeptidase A