Arg(1098) is critical for the chloride dependence of human angiotensin I-converting enzyme C-domain catalytic activity

• Published in: The Journal of biological chemistry 2001-09, Vol.276 (36), p.33518
• Main Authors: Liu, X, Fernandez, M, Wouters, M A, Heyberger, S, Husain, A
• Format: Article
• Language: English
• Subjects: Allosteric Site; Amino Acid Sequence; Animals; Arginine - chemistry; Binding Sites; Catalysis; Catalytic Domain; Chlorine - chemistry; Chlorine - physiology; COS Cells; Dose-Response Relationship, Drug; Enzyme Activation; Glutamine - chemistry; Humans; Kinetics; Molecular Sequence Data; Peptides - chemistry; Peptidyl-Dipeptidase A - chemistry; Protein Binding; Protein Structure, Tertiary; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Sequence Homology, Amino Acid; Sodium Chloride - pharmacology; Transfection
• ISSN: 0021-9258
• DOI: 10.1074/jbc.M101495200

Angiotensin (Ang) I-converting enzyme (ACE) is a Zn(2+) metalloprotease with two homologous catalytic domains. Both the N- and C-terminal domains are peptidyl dipeptidases. Hydrolysis by ACE of its decapeptide substrate Ang I is increased by Cl(-), but the molecular mechanism of this regulation is unclear. A search for single substitutions to Gln among all conserved basic residues (Lys/Arg) in human ACE C-domain identified R1098Q as the sole mutant that lacked Cl(-) dependence. Cl(-) dependence is also lost when the equivalent Arg in the N-domain, Arg(500), is substituted with Gln. The Arg(1098) to Lys substitution reduced Cl(-) binding affinity by approximately 100-fold. In the absence of Cl(-), substrate binding affinity (1/K(m)) of and catalytic efficiency (k(cat)/K(m)) for Ang I hydrolysis are increased 6.9- and 32-fold, respectively, by the Arg(1098) to Gln substitution, and are similar (