A novel angiotensin I-converting enzyme mutation (S333W) impairs N-domain enzymatic cleavage of the anti-fibrotic peptide, AcSDKP.

BACKGROUND: Angiotensin I-converting enzyme (ACE) has two functional N- and C-domain active centers that display differences in the metabolism of biologically-active peptides including the hemoregulatory tetrapeptide, Ac-SDKP, hydrolysed preferentially by the N domain active center. Elevated Ac-SDKP concentrations are associated with reduced tissue fibrosis. RESULTS: We identified a patient of African descent exhibiting unusual blood ACE kinetics with reduced relative hydrolysis of two synthetic ACE substrates (ZPHL/HHL ratio) suggestive of the ACE N domain center inactivation. Inhibition of blood ACE activity by anti-catalytic mAbs and ACE inhibitors and conformational fingerprint of blood ACE suggested overall conformational changes in the ACE molecule and sequencing identified Ser333Trp substitution in the N domain of ACE. In silico analysis demonstrated S333W localized in the S1 pocket of the active site of the N domain with the bulky Trp adversely affecting binding of ACE substrates due to steric hindrance. Expression of mutant ACE (S333W) in CHO cells confirmed altered kinetic properties of mutant ACE and conformational changes in the N domain. Further, the S333W mutant displayed decreased ability (5-fold) to cleave the physiological substrate AcSDKP compared to wild-type ACE. CONCLUSIONS AND SIGNIFICANCE: A novel Ser333Trp ACE mutation results in dramatic changes in ACE kinetic properties and lowered clearance of Ac-SDKP. Individuals with this mutation (likely with significantly increased levels of the hemoregulatory tetrapeptide in blood and tissues), may confer protection against fibrosis.

Angiotensin I-converting enzyme mutation (Trp1197Stop) causes a dramatic increase in blood ACE.

BACKGROUND: Angiotensin-converting enzyme (ACE) metabolizes many peptides and plays a key role in blood pressure regulation and vascular remodeling. Elevated ACE levels may be associated with an increased risk for different cardiovascular or respiratory diseases, including asthma. Previously, a molecular mechanism underlying a 5-fold familial increase of blood ACE was discovered: Pro1199Leu substitution enhanced the cleavage-secretion process. Carriers of this mutation were Caucasians from Europe (mostly Dutch) or had European roots. METHODOLOGY/PRINCIPAL FINDINGS: We have found a family of African-American descent whose affected members' blood ACE level was increased 13-fold over normal. In affected family members, codon TGG coding for Trp1197 was substituted in one allele by TGA (stop codon). As a result, half of ACE expressed in these individuals had a length of 1196 amino acids and lacked a transmembrane anchor. This ACE mutant is not trafficked to the cell membrane and is directly secreted out of cells; this mechanism apparently accounts for the high serum ACE level seen in affected individuals. A haplotype of the mutant ACE allele was determined based on 12 polymorphisms, which may help to identify other carriers of this mutation. Some but not all carriers of this mutation demonstrated airflow obstruction, and some but not all have hypertension. CONCLUSIONS/SIGNIFICANCE: We have identified a novel Trp1197Stop mutation that results in dramatic elevation of serum ACE. Since blood ACE elevation is often taken as a marker of disease activity (sarcoidosis and Gaucher diseases), it is important for clinicians and medical scientists to be aware of alternative genetic causes of elevated blood ACE that are not apparently linked to disease.