title: Angiotensin I-converting enzyme Gln1069Arg mutation impairs trafficking to the cell surface resulting in selective denaturation of the C-domain.
abstract: BACKGROUND
  Angiotensin-converting enzyme (ACE; Kininase II; CD143) hydrolyzes small peptides such as angiotensin I, bradykinin, substance P, LH-RH and several others and thus plays a key role in blood pressure regulation and vascular remodeling. Complete absence of ACE in humans leads to renal tubular dysgenesis (RTD), a severe disorder of renal tubule development characterized by persistent fetal anuria and perinatal death.
  
  
  RESULTS
  Patient with RTD in Lisbon, Portugal, maintained by peritoneal dialysis since birth, was found to have a homozygous substitution of Arg for Glu at position 1069 in the C-terminal domain of ACE (Q1069R) resulting in absence of plasma ACE activity; both parents and a brother who are heterozygous carriers of this mutation had exactly half-normal plasma ACE activity compared to healthy individuals. We hypothesized that the Q1069R substitution impaired ACE trafficking to the cell surface and led to accumulation of catalytically inactive ACE in the cell cytoplasm. CHO cells expressing wild-type (WT) vs. Q1069R-ACE demonstrated the mutant accumulates intracellularly and also that it is significantly degraded by intracellular proteases. Q1069R-ACE retained catalytic and immunological characteristics of WT-ACE N domain whereas it had 10-20% of the nativity of the WT-ACE C domain. A combination of chemical (sodium butyrate) or pharmacological (ACE inhibitor) chaperones with proteasome inhibitors (MG 132 or bortezomib) significantly restored trafficking of Q1069R-ACE to the cell surface and increased ACE activity in the cell culture media 4-fold.
  
  
  CONCLUSIONS
  Homozygous Q1069R substitution results in an ACE trafficking and processing defect which can be rescued, at least in cell culture, by a combination of chaperones and proteasome inhibitors. Further studies are required to determine whether similar treatment of individuals with this ACE mutation would provide therapeutic benefits such as concentration of primary urine.
authors.affiliation: Department of Anesthesiology, University of Illinois at Chicago, Chicago, Illinois, United States of America. danilov@uic.edu
authors.forename: Sergei M
authors.initials: SM
authors.lastname: Danilov
authors.forename: Sergey
authors.initials: S
authors.lastname: Kalinin
authors.forename: Zhenlong
authors.initials: Z
authors.lastname: Chen
authors.forename: Elena I
authors.initials: EI
authors.lastname: Vinokour
authors.forename: Andrew B
authors.initials: AB
authors.lastname: Nesterovitch
authors.forename: David E
authors.initials: DE
authors.lastname: Schwartz
authors.forename: Olivier
authors.initials: O
authors.lastname: Gribouval
authors.forename: Marie-Claire
authors.initials: MC
authors.lastname: Gubler
authors.forename: Richard D
authors.initials: RD
authors.lastname: Minshall
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D055162:Biocatalysis / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D005190:Family; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D008297:Male; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000276:immunology; D009154:Mutation / Q000235:genetics*; D007703:Peptidyl-Dipeptidase A / Q000097:blood / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects; D011994:Recombinant Proteins / Q000235:genetics; D013696:Temperature
chemical_list: D018832:Molecular Chaperones; D050505:Mutant Proteins; D011994:Recombinant Proteins; D007703:Peptidyl-Dipeptidase A
references.citation: Tissue Antigens. 2010 Feb;75(2):136-50
references.pmid: 20003136
references.citation: Biochemistry. 2006 Oct 24;45(42):12654-63
references.pmid: 17042482
references.citation: Circ Res. 2003 Jul 25;93(2):148-54
references.pmid: 12805239
references.citation: Protein Expr Purif. 1991 Feb;2(1):1-9
references.pmid: 1668266
references.citation: Eur J Pediatr. 2009 Feb;168(2):207-9
references.pmid: 18478260
references.citation: J Proteome Res. 2008 Aug;7(8):3396-411
references.pmid: 18576678
references.citation: Tissue Antigens. 2008 Oct;72(4):354-68
references.pmid: 18700874
references.citation: Nature. 2003 Jan 30;421(6922):551-4
references.pmid: 12540854
references.citation: Biochem J. 2003 Apr 15;371(Pt 2):437-42
references.pmid: 12542396
references.citation: J Pharmacol Exp Ther. 1996 Dec;279(3):1582-9
references.pmid: 8968386
references.citation: J Biol Chem. 1991 May 15;266(14):9002-8
references.pmid: 1851160
references.citation: Biochemistry. 2007 Aug 7;46(31):9019-31
references.pmid: 17630779
references.citation: Regul Pept. 1995 Nov 10;59(3):379-87
references.pmid: 8577943
references.citation: PLoS One. 2009 Dec 14;4(12):e8282
references.pmid: 20011602
references.citation: Biochem Pharmacol. 1996 Jan 12;51(1):11-4
references.pmid: 8534262
references.citation: Biol Chem. 2008 Sep;389(9):1153-61
references.pmid: 18713002
references.citation: J Biol Chem. 1996 Mar 15;271(11):6429-34
references.pmid: 8626443
references.citation: J Mol Biol. 2006 Mar 31;357(3):964-74
references.pmid: 16476442
references.citation: Clin Chem. 2005 Jun;51(6):1040-3
references.pmid: 15914791
references.citation: Nephrol Dial Transplant. 1998 Dec;13(12):2991-4
references.pmid: 9870448
references.citation: J Proteome Res. 2007 Apr;6(4):1580-94
references.pmid: 17326675
references.citation: Exp Hematol. 2003 Dec;31(12):1301-9
references.pmid: 14662338
references.citation: J Proteome Res. 2005 Mar-Apr;4(2):258-67
references.pmid: 15822901
references.citation: Histochemistry. 1986;85(1):73-80
references.pmid: 3015846
references.citation: In Vitro Cell Dev Biol Anim. 1998 Jul-Aug;34(7):545-54
references.pmid: 9719414
references.citation: J Biol Chem. 1994 Oct 28;269(43):26806-14
references.pmid: 7523412
references.citation: J Am Soc Nephrol. 1993 Mar;3(9):1575-82
references.pmid: 8507813
references.citation: J Mol Recognit. 1995 Jan-Apr;8(1-2):72-6
references.pmid: 7541231
references.citation: J Hypertens. 1997 Dec;15(12 Pt 2):1579-92
references.pmid: 9488209
references.citation: Neurochem Int. 2003 Jul;43(1):1-7
references.pmid: 12605877
references.citation: Curr Opin Struct Biol. 2000 Aug;10(4):438-42
references.pmid: 10981632
references.citation: Nature. 1995 May 11;375(6527):146-8
references.pmid: 7753170
references.citation: Biochemistry. 1991 Jul 23;30(29):7118-26
references.pmid: 1649623
references.citation: Eur Heart J. 2002 Jul;23(14):1083-6
references.pmid: 12090745
references.citation: Biochim Biophys Acta. 1970 Apr 22;206(1):136-42
references.pmid: 4315054
references.citation: Hybridoma (Larchmt). 2005 Feb;24(1):14-26
references.pmid: 15785205
references.citation: Am J Physiol. 1999 May;276(5):F751-7
references.pmid: 10330057
references.citation: Hypertension. 2000 Jul;36(1):116-21
references.pmid: 10904022
references.citation: Arch Biochem Biophys. 1983 Nov;227(1):188-201
references.pmid: 6314908
references.citation: Biol Chem. 2006 Aug;387(8):1043-51
references.pmid: 16895474
references.citation: Neuroendocrinology. 1983 Jul;37(1):32-40
references.pmid: 6310427
references.citation: Circ Res. 2005 Jun 10;96(11):1135-44
references.pmid: 15947253
references.citation: N Engl J Med. 2005 May 12;352(19):1992-2001
references.pmid: 15888700
references.citation: Nat Genet. 2005 Sep;37(9):964-8
references.pmid: 16116425
references.citation: Biochemistry. 2008 Jun 3;47(22):5942-50
references.pmid: 18457420
references.citation: J Am Soc Nephrol. 2006 Aug;17(8):2253-63
references.pmid: 16790508
references.citation: J Cardiovasc Pharmacol. 2008 Jul;52(1):90-103
references.pmid: 18645413
references.citation: N Engl J Med. 2006 Jun 8;354(23):2443-51
references.pmid: 16760444
references.citation: Expert Opin Investig Drugs. 2007 Jun;16(6):819-28
references.pmid: 17501694
references.citation: Biochemistry. 2006 Apr 18;45(15):4831-47
references.pmid: 16605251
references.citation: Tissue Antigens. 2005 Mar;65(3):240-51
references.pmid: 15730516
references.citation: Biochem J. 1987 Feb 1;241(3):625-33
references.pmid: 2439065
references.citation: Tissue Antigens. 2003 Jan;61(1):49-62
references.pmid: 12622775
references.citation: Biochemistry. 1989 Jun 27;28(13):5311-8
references.pmid: 2476171
references.citation: Early Hum Dev. 2006 Jan;82(1):23-8
references.pmid: 16427219
references.citation: Science. 1996 Sep 20;273(5282):1725-8
references.pmid: 8781238
references.citation: J Cell Biol. 2000 Sep 4;150(5):1057-70
references.pmid: 10973995
references.citation: J Biol Chem. 1993 May 5;268(13):9496-503
references.pmid: 7683654
references.citation: Biochem J. 2005 Nov 1;391(Pt 3):641-7
references.pmid: 16033330
references.citation: Am J Clin Pathol. 1976 Aug;66(2):416-24
references.pmid: 181981
references.citation: Science. 1976 Mar 12;191(4231):1050-1
references.pmid: 175444
references.citation: Biochemistry. 1996 Jul 23;35(29):9560-6
references.pmid: 8755737
references.citation: FEBS Lett. 2002 Jul 3;522(1-3):77-82
references.pmid: 12095622
references.citation: Nature. 2003 Dec 18;426(6968):905-9
references.pmid: 14685252
references.citation: Hum Mol Genet. 2008 Oct 15;17(20):3180-90
references.pmid: 18647753
references.citation: Traffic. 2004 Nov;5(11):821-37
references.pmid: 15479448
references.citation: Proc Natl Acad Sci U S A. 1988 Dec;85(24):9386-90
references.pmid: 2849100
references.citation: Biochemistry (Mosc). 2004 Nov;69(11):1233-8
references.pmid: 15627377
references.citation: Nat Rev Cancer. 2004 May;4(5):349-60
references.pmid: 15122206
references.citation: Isr J Med Sci. 1978 Mar;14(3):314-8
references.pmid: 205523