Angiotensin I-converting enzyme Gln1069Arg mutation impairs trafficking to the cell surface resulting in selective denaturation of the C-domain.
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.
Danilov
Sergei M
SM
Department of Anesthesiology, University of Illinois at Chicago, Chicago, Illinois, United States of America. danilov@uic.edu
Kalinin
Sergey
S
Chen
Zhenlong
Z
Vinokour
Elena I
EI
Nesterovitch
Andrew B
AB
Schwartz
David E
DE
Gribouval
Olivier
O
Gubler
Marie-Claire
MC
Minshall
Richard D
RD
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
D018832:Molecular Chaperones; D050505:Mutant Proteins; D011994:Recombinant Proteins; D007703:Peptidyl-Dipeptidase A
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