Surface Plasmon Resonance Analysis of the Binding Mechanism of Pharmacological and Peptidic Inhibitors to Human Somatic Angiotensin I-Converting Enzyme

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.

Lysozyme and bilirubin bind to ACE and regulate its conformation and shedding

“…ACE interaction with lysozyme, a well-studied protein, was unanticipated and had not previously been reported possibly reflecting weak binding constant of ACE-lysozyme interaction with very high association and dissociation rates of ACE-lysozyme complex ( Fig. S5B ) observed for ACE-caseinopeptide interactions 33 . Conversely, this may reflect the use of testicular ACE for co-immunoprecipitation in prior studies 14 15 17 , which does not have a cleft between domains.…”

Revealing the ACE receptor binding properties and interaction mechanisms of salty oligopeptides from Stropharia rugosoannulata mushroom by molecular simulation and antihypertensive evaluation

“…Comparing the bioinformatics computer-assisted analyses with the target protein-peptide molecular interaction analyses, obtaining a comprehensive understanding of the interacting characteristics through cross-validation of multiple techniques, it is of great reference significance for elucidating the receptor binding properties and interaction mechanism of S. rugosoannulata ACE inhibitory oligopeptides. The primary methods for analyzing the interaction mechanism of ACE inhibitory peptides based on biomolecular interactions include fluorescence spectroscopy, 18,19,26,[28][29][30] surface plasmon resonance, 31,32 circular dichroism, [18][19][20][28][29][30]33 isothermal titration calorimetry, 19,30,33,34 and so on. ITC assay demonstrated a 1 : 1 stoichiometry between lisinopril, captopril, enalaprilat, and ACE.…”

“…The primary methods for analyzing the interaction mechanism of ACE inhibitory peptides based on biomolecular interactions include fluorescence spectroscopy, 18,19,26,28–30 surface plasmon resonance, 31,32 circular dichroism, 18–20,28–30,33 isothermal titration calorimetry, 19,30,33,34 and so on. ITC assay demonstrated a 1 : 1 stoichiometry between lisinopril, captopril, enalaprilat, and ACE.…”

“…The ACE inhibitory activity of ANSEVAQWR was more potent than that of EALVSQLTR, with IC 50 values of 89.58 and 91.48 μM, respectively. Zidane and his colleagues 32 conducted an SPR study to examine how ACE interacted with two bovine peptidic inhibitors from bovine α s2-casein, namely ‘F 174 ALPQYLK 181’ and ‘F 174 ALPQY 179’. Both inhibitors showed an IC 50 of 4.3 μM.…”