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An angiotensin I-converting enzyme mutation (Y465D) causes a dramatic increase in blood ACE via accelerated ACE shedding.
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Recently, we found a family whose affected members had a 6-fold increase in blood ACE and a Tyr465Asp (Y465D) substitution, distal to the stalk region, in the N domain of ACE.The Y465D substitution results in dramatic increase in the rate of ACE shedding and is associated with significant local conformational changes in ACE.These changes could result in increased ACE dimerization and accessibility of the stalk region or the entire sACE, thus increasing the rate of cleavage by the putative ACE secretase (sheddase).
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PubMed Central - PubMed
Affiliation: Department of Anesthesiology, University of Illinois at Chicago, Chicago, Illinois, United States of America. danilov@uic.edu
ABSTRACT
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Background: Angiotensin I-converting enzyme (ACE) metabolizes a range of peptidic substrates and plays a key role in blood pressure regulation and vascular remodeling. Thus, elevated ACE levels may be associated with an increased risk for different cardiovascular or respiratory diseases. Previously, a striking familial elevation in blood ACE was explained by mutations in the ACE juxtamembrane region that enhanced the cleavage-secretion process. Recently, we found a family whose affected members had a 6-fold increase in blood ACE and a Tyr465Asp (Y465D) substitution, distal to the stalk region, in the N domain of ACE. Methodology/principal findings: HEK and CHO cells expressing mutant (Tyr465Asp) ACE demonstrate a 3- and 8-fold increase, respectively, in the rate of ACE shedding compared to wild-type ACE. Conformational fingerprinting of mutant ACE demonstrated dramatic changes in ACE conformation in several different epitopes of ACE. Cell ELISA carried out on CHO-ACE cells also demonstrated significant changes in local ACE conformation, particularly proximal to the stalk region. However, the cleavage site of the mutant ACE--between Arg1203 and Ser1204--was the same as that of WT ACE. The Y465D substitution is localized in the interface of the N-domain dimer (from the crystal structure) and abolishes a hydrogen bond between Tyr465 in one monomer and Asp462 in another. Conclusions/significance: The Y465D substitution results in dramatic increase in the rate of ACE shedding and is associated with significant local conformational changes in ACE. These changes could result in increased ACE dimerization and accessibility of the stalk region or the entire sACE, thus increasing the rate of cleavage by the putative ACE secretase (sheddase). Related in: MedlinePlus |
![Epitope mapping of human sACE using EM-based model.Model of human sACE was generated using an EM-based model of porcine sACE [49]. Epitopes localization (colored spots) were taken from the following publications [40], [51]–[55].. Relevant amino acid residues in the ectodomain and juxtamembrane region of ACE and a cartoon of the membrane-bound ACE secretase are indicated: Leu1 and Asp616-N terminal residues of the N- and C-domains, respectively; Pro1193 –the last amino acid residues seen in the 3D structure of C domain; Pro1199 – localization of P1199L mutation causing 5-fold increase in blood ACE levels [32]; Arg1203-C-terminal amino acid residue of soluble ACE [23]; Val1228-Ser1248-transmembrane domain [65]; Ser1277-C-terminal amino acid residue of full-length somatic ACE [17].](/imgs/512/278/3187827/PMC3187827_pone.0025952.g005.png?keywords=spots)
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pone-0025952-g005: Epitope mapping of human sACE using EM-based model.Model of human sACE was generated using an EM-based model of porcine sACE [49]. Epitopes localization (colored spots) were taken from the following publications [40], [51]–[55].. Relevant amino acid residues in the ectodomain and juxtamembrane region of ACE and a cartoon of the membrane-bound ACE secretase are indicated: Leu1 and Asp616-N terminal residues of the N- and C-domains, respectively; Pro1193 –the last amino acid residues seen in the 3D structure of C domain; Pro1199 – localization of P1199L mutation causing 5-fold increase in blood ACE levels [32]; Arg1203-C-terminal amino acid residue of soluble ACE [23]; Val1228-Ser1248-transmembrane domain [65]; Ser1277-C-terminal amino acid residue of full-length somatic ACE [17]. Mentions: There was a consistent loss of affinity for the N-domain specific mAb 5F1 with cell-associated and soluble mutant ACE in the precipitation assay (Fig. 3) as well as membrane-bound ACE-Y465D in the cell ELISA (Fig. 4B). Y465 might participate directly in mAb 5F1 binding to ACE, because it is localized close to/within the epitope for mAb 5F1 [53] (Fig. 5), or the mutation has resulted in gross conformational changes that have altered the mAb 5F1 epitope. Alternatively, the changes in putative ACE dimerization have masked the epitope for mAb 5F1. |
![Epitope mapping of human sACE using EM-based model.Model of human sACE was generated using an EM-based model of porcine sACE [49]. Epitopes localization (colored spots) were taken from the following publications [40], [51]–[55].. Relevant amino acid residues in the ectodomain and juxtamembrane region of ACE and a cartoon of the membrane-bound ACE secretase are indicated: Leu1 and Asp616-N terminal residues of the N- and C-domains, respectively; Pro1193 –the last amino acid residues seen in the 3D structure of C domain; Pro1199 – localization of P1199L mutation causing 5-fold increase in blood ACE levels [32]; Arg1203-C-terminal amino acid residue of soluble ACE [23]; Val1228-Ser1248-transmembrane domain [65]; Ser1277-C-terminal amino acid residue of full-length somatic ACE [17].](/imgs/512/278/3187827/PMC3187827_pone.0025952.g005.png)
View Article: PubMed Central - PubMed
Affiliation: Department of Anesthesiology, University of Illinois at Chicago, Chicago, Illinois, United States of America. danilov@uic.edu
Background: Angiotensin I-converting enzyme (ACE) metabolizes a range of peptidic substrates and plays a key role in blood pressure regulation and vascular remodeling. Thus, elevated ACE levels may be associated with an increased risk for different cardiovascular or respiratory diseases. Previously, a striking familial elevation in blood ACE was explained by mutations in the ACE juxtamembrane region that enhanced the cleavage-secretion process. Recently, we found a family whose affected members had a 6-fold increase in blood ACE and a Tyr465Asp (Y465D) substitution, distal to the stalk region, in the N domain of ACE.
Methodology/principal findings: HEK and CHO cells expressing mutant (Tyr465Asp) ACE demonstrate a 3- and 8-fold increase, respectively, in the rate of ACE shedding compared to wild-type ACE. Conformational fingerprinting of mutant ACE demonstrated dramatic changes in ACE conformation in several different epitopes of ACE. Cell ELISA carried out on CHO-ACE cells also demonstrated significant changes in local ACE conformation, particularly proximal to the stalk region. However, the cleavage site of the mutant ACE--between Arg1203 and Ser1204--was the same as that of WT ACE. The Y465D substitution is localized in the interface of the N-domain dimer (from the crystal structure) and abolishes a hydrogen bond between Tyr465 in one monomer and Asp462 in another.
Conclusions/significance: The Y465D substitution results in dramatic increase in the rate of ACE shedding and is associated with significant local conformational changes in ACE. These changes could result in increased ACE dimerization and accessibility of the stalk region or the entire sACE, thus increasing the rate of cleavage by the putative ACE secretase (sheddase).
