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Surface plasmon resonance analysis of the binding mechanism of pharmacological and peptidic inhibitors to human somatic angiotensin I-converting enzyme.

Biochemistry (2013-10-31)
Faïza Zidane, Gabrielle Zeder-Lutz, Danièle Altschuh, Jean-Michel Girardet, Laurent Miclo, Catherine Corbier, Céline Cakir-Kiefer
ABSTRACT

Somatic angiotensin I-converting enzyme (ACE) possesses two catalytic domains and plays a major role in the regulation of blood pressure, thus representing a therapeutic target for the treatment of hypertension. We present a comprehensive surface plasmon resonance (SPR) study of the interaction of human somatic ACE with the pharmacological inhibitors captopril and lisinopril, the bradykinin potentiating peptide BPP-11b, and the food peptidic inhibitors from bovine αs2-casein, F(174)ALPQYLK(181) and F(174)ALPQY(179). SPR binding curves recorded with the high potency inhibitors captopril, lisinopril, and BPP-11b were evaluated both by regression analysis and by kinetic distribution analysis. The results indicated that captopril and lisinopril bound ACE with two K(D)'s differing by a factor 10-20 and >30, respectively (lowest K(D) = 0.1-0.3 nM for both inhibitors). This shows, for the first time in a direct binding assay with the two-domain enzyme, the existence of two binding modes of the pharmacological inhibitors, presumably with the two ACE domains. The BPP-11b-ACE binding curves were complex but showed a predominant interaction with K(D) in the nanomolar range. The caseinopeptides, known to inhibit ACE with an IC₅₀ of 4.3 μM, bound to ACE with K(D) = 3-4 μM. Mapping of the F(174)ALPQY(179) binding site on ACE by sequential binding studies using captopril or BPP-11b indicated that it bound to (or near) the two active sites of ACE, in agreement with the stoichiometry of 2 determined from data fitting. Our results provide a detailed characterization of ACE-inhibitor binding modes and validate SPR for predicting the inhibitory potential of new compounds.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Captopril, meets USP testing specifications
Sigma-Aldrich
Captopril, ≥98% (HPLC), powder
Supelco
Captopril, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Lisinopril, ≥98% (HPLC)
Supelco
Lisinopril, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Angiotensin Converting Enzyme from rabbit lung, ≥2.0 units/mg protein (modified Warburg-Christian)
Sigma-Aldrich
Angiotensin Converting Enzyme from porcine kidney, lyophilized powder, ≥10 units/mg protein (Bradford)