In vitro and in silico studies of novel synthetic ACE-inhibitory peptides derived from Saccharomyces cerevisiae protein hydrolysate

The structure-function relation of YR-10 (YGKPVAVPAR) was investigated by synthesizing four structural analogs of that including YHR-10 (YGKHVAVHAR), GA-8 (GKPVAVPA), GHA-8 (GKHVAVHA), and PAR-3 (PAR). GA-8 (GKPVAVPA) was synthesized on the basis of simulated enzymatic gastrointestinal digestion performed by bioinformatics tools (expasy-peptide cutter). This study explains the molecular mechanisms for the interaction of synthetic peptides with ACE. The IC₅₀ values of each were 139.554 ± 2.3, 61.91 ± 1.2, 463.230 ± 3.56, 135.135 ± 2.1, 514.024 ± 5.86 µM, respectively. Results indicated that Pro replacement with His in YR-10 and GA-8 increased ACE inhibitory activity respectively, by 55.63% and 70.82%. Removal of Tyr and Arg from respectively N and C terminal positions of YR-10, following in silico simulated gastrointestinal digestion caused the 3.31 fold decrease in ACE inhibitory activity. YHR-10 showed the best docking poses, and GHA-8 exhibited interaction with Zn²⁺. Lineweaver–Burk plots of most active peptides suggest that they act as noncompetitive inhibitors against ACE.     

Sequences of tryptic peptides containing the five cysteinyl residues of ribulosebisphosphate carboxylase/oxygenase from Rhodospirillum rubrum

Tryptic peptides which account for all five cysteinyl residues in ribulosebisphosphate carboxylase/oxygenase from Rhodospirillum rubrum have been purified and sequenced. Collectively, these peptides contain 94 of the approximately 500 amino acid residues per molecule of subunit. Due to one incomplete cleavage at a site for trypsin and two incomplete chymotryptic-like cleavages, eight major radioactive peptides (rather than five as predicted) were recovered from tryptic digests of the enzyme that had been carboxymethylated with [³H]iodoacetate. The established sequences are: GlyTyrThrAlaPheValHisCys¹Lys TyrValAspLeuAlaLeuLysGluGluAspLeuIleAla GlyGlyGluHisValLeuCys¹AlaTyr AlaGlyTyrGlyTyrValAlaThrAlaAlaHisPheAla AlaGluSerSerThrGlyThrAspValGluValCys¹ ThrThrAsxAsxPheThrArg AlaCys¹ThrProIleIleSerGlyGlyMetAsnAla LeuArg ProPheAlaGluAlaCys¹HisAlaPheTrpLeuGly GlyAsnPheIleLys In these peptides, radioactive carboxymethylcysteinyl residues are denoted with asterisks and the sites of incomplete cleavage with vertical wavy lines. None of the peptides appear homologous with either of two cysteinyl-containing, active-site peptides previously isolated from spinach ribulosebisphosphate carboxylase/oxygenase.     

Characterization of the Particles of Purified κ-Casein: Trypsin as a Probe of Surface-Accessible Residues

κ-Casein as purified from bovine milk exhibits a rather unique disulfide bonding pattern as revealed by SDS–PAGE. The disulfide-bonded caseins present range from dimer to octamer and above and preparations contain about 10% monomer. All of these heterogeneous polymers, however, self-associate into nearly spherical particles with an average diameter of 13 nm at pH 8.0, as revealed by negatively stained transmission electron micrographs and dynamic light scattering. The weight-average molecular weight of the aggregates at pH 8.0, as judged by analytical ultracentrifugation, is 648,000. Trypsin digestion at pH 8.0 was used to probe the surface groups of the κ-casein A polymers. The reaction with trypsin was rapid and the peptides liberated were identified by separation with reverse-phase HPLC, amino acid analysis, and protein sequencing. The most rapidly released peptides (t _1/2 < 30 sec) were from cleavage at Arg 97 and Lys residues 111 and 112. These results suggest a surface orientation for these residues, and the data are in accord with earlier proposed 3D predictive models for κ-casein. It is speculated that Arg 97, together with adjacent His residues (98 and 100) and Lys residues 111 and 112, form two positively charged clusters on the surface of the otherwise negatively charged casein. These clusters bracket the neutral chymosin cleavage site (whose hydrolysis triggers a well-known digestive process) and so these clusters may facilitate docking of the substrate caseins with chymosin.     

Molecular Dynamics Simulation and Molecular Docking Studies of Angiotensin Converting Enzyme with Inhibitor Lisinopril and Amyloid Beta Peptide

Angiotensin converting enzyme (ACE) cleaves amyloid beta peptide. So far this cleavage mechanism has not been studied in detail at atomic level. Keeping this view in mind, we performed molecular dynamics simulation of crystal structure complex of testis truncated version of ACE (tACE) and its inhibitor lisinopril along with Zn²⁺ to understand the dynamic behavior of active site residues of tACE. Root mean square deviation results revealed the stability of tACE throughout simulation. The residues Ala 354, Glu 376, Asp 377, Glu 384, His 513, Tyr 520 and Tyr 523 of tACE stabilized lisinopril by hydrogen bonding interactions. Using this information in subsequent part of study, molecular docking of tACE crystal structure with Aβ-peptide has been made to investigate the interactions of Aβ-peptide with enzyme tACE. The residues Asp 7 and Ser 8 of Aβ-peptide were found in close contact with Glu 384 of tACE along with Zn²⁺. This study has demonstrated that the residue Glu 384 of tACE might play key role in the degradation of Aβ-peptide by cleaving peptide bond between Asp 7 and Ser 8 residues. Molecular basis generated by this attempt could provide valuable information towards designing of new therapies to control Aβ concentration in Alzheimer’s patient.     

In silico identification of novel ACE and DPP-IV inhibitory peptides derived from buffalo milk proteins and evaluation of their inhibitory mechanisms

The capacity of buffalo milk proteins to release bioactive peptides was evaluated and novel bioactive peptides were identified. The sequential similarity between buffalo milk proteins and their cow counterparts was analysed. Buffalo milk proteins were simulated to yield theoretical peptides via in silico proteolysis. The potential of selected proteins to release specific bioactive peptides was evaluated by the A value obtained from the BIOPEP–UWM database (Minkiewicz et al. in Int J Mol Sci 20(23):5978, 2019). Buffalo milk protein is a suitable precursor to produce bioactive peptides, particularly dipeptidyl peptidase IV (DPP-IV) and angiotensin I-converting enzyme (ACE) inhibitory peptides. Two novel ACE inhibitory peptides (KPW and RGP) and four potential DPP-IV inhibitory peptides (RGP, KPW, FPK and KFTW) derived from in silico proteolysis of buffalo milk proteins were screened using different integrated bioinformatic approaches (PeptideRanker, Innovagen, peptide-cutter and molecular docking). The Lineweaver–Burk plots showed that KPW (IC₅₀?=?136.28?±?10.77 μM) and RGP (104.72?±?8.37 μM) acted as a competitive inhibitor against ACE. Similarly, KFTW (IC₅₀?=?873.92?±?32.89 μM) was also a competitive inhibitor of DPP-IV, while KPW and FPK (82.52?±?10.37 and 126.57?±?8.45 μM, respectively) were mixed-type inhibitors. It should be emphasized that this study does not involve any clinical trial.

     

In silico study of carvone derivatives as potential neuraminidase inhibitors

Recent outbreaks of highly pathogenic influenza strains have highlighted the need to develop new anti-influenza drugs. Here, we report an in silico study of carvone derivatives to analyze their binding modes with neuraminidase (NA) active sites. Two proposed carvone analogues, CV(A) and CV(B), with 36 designed ligands were predicted to inhibit NA (PDB ID: 3TI6) using molecular docking. The design is based on structural resemblance with the commercial inhibitor, oseltamivir (OTV), ligand polarity, and amino acid residues in the NA active sites. Docking simulations revealed that ligand A18 has the lowest energy binding (?G_bind) value of ?8.30 kcal mol^-1, comparable to OTV with ?G_bind of ?8.72 kcal mol^-1. A18 formed seven hydrogen bonds (H-bonds) at residues Arg292, Arg371, Asp151, Trp178, Glu227, and Tyr406, while eight H-bonds were formed by OTV with amino acids Arg118, Arg292, Arg371, Glu119, Asp151, and Arg152. Molecular dynamics (MD) simulation was conducted to compare the stability between ligand A18 and OTV with NA. Our simulation study showed that the A18-NA complex is as stable as the OTV-NA complex during the MD simulation of 50 ns through the analysis of RMSD, RMSF, total energy, hydrogen bonding, and MM/PBSA free energy calculations.     

Amino acid sequence of seminalplasmin, an antimicrobial protein from bull semen

Analytical ultracentrifugation of highly purified seminalplasmin revealed a molecular mass of 6300. Amino acid analysis of the protein preparation indicated the absence of sulfur-containing amino acids cysteine and methionine. The amino acid sequence of seminalplasmin was determined by manual Edman degradation of peptides obtained by proteolytic enzymes trypsin, chymotrypsin and thermolysin: NH₂-Ser Asp Glu Lys Ala Ser Pro Asp Lys His His Arg Phe Ser Leu Ser Arg Tyr Ala Lys Leu Ala Asn Arg Leu Ser Lys Trp Ile Gly Asn Arg Gly Asn Arg Leu Ala Asn Pro Lys Leu Leu Glu Thr Phe Lys Ser Val-COOH. The number of amino acids according to the sequence were 48, the molecular mass 6385. As predicted from the sequence, seminalplasmin very likely contains two α-helical domains in which residues 8-17 and 40-48 are involved. No evidence for the existence of β-sheet structures was obtained. Treatment of seminalplasmin with the above proteases as well as with amino peptidase M and carboxypeptidase Y completely eliminated biological activity.     

Purification and identification of novel angiotensin-I converting enzyme (ACE) inhibitory peptides from cultured marine microalgae (Nannochloropsis oculata) protein hydrolysate

Isolation of bioactive compounds and commercialization of marine microalgae sources are interesting targets in future marine biotechnology. Cultured biomass of the marine microalga, Nannochloropsis oculata, was used to purify angiotensin-I converting enzyme (ACE) inhibitory peptides using proteases including pepsin, trypsin, α-chymotrypsin, papain, alcalase, and neutrase. The pepsin hydrolysate exhibited the highest ACE inhibitory activity, compared to the other hydrolysates and then was separated into three fractions (F1, F2, and F3) using Sephadex G-25 gel filtration column chromatography. First fraction (F1) showed the highest ACE inhibitory activity and it was further purified into two fractions (F1-1 and F1-2) using reverse-phase high-performance liquid chromatography. The IC₅₀ value of purified ACE inhibitory peptides were 123 and 173 μM and identified as novel peptides, Gly-Met-Asn-Asn-Leu-Thr-Pro (GMNNLTP; MW, 728 Da) and Leu-Glu-Gln (LEQ; MW, 369 Da), respectively. In addition, nitric oxide production level (%) was significantly increased by the purified peptide (Gly-Met-Asn-Asn-Leu-Thr-Pro) compared to the purified peptide (Leu-Glu-Gln) and other treated pepsin hydrolysate fractions on human umbilical vein endothelial cells (HUVECs). Cell viability assay showed no cytotoxicity on HUVECs with the treated purified peptides and fractions. These results suggest that the isolated peptides from cultured marine microalga, N. oculata protein sources may have potentiality to use commercially as ACE inhibitory agents in functional food industry.     

Bioactive Peptides from Bovine Milk α-Casein: Isolation,Characterization and Multifunctional properties

α-Casein group of proteins makes up to 65% of the total casein and consists of α_S1- casein_, α_S2- casein and other related proteins. Among all the proteases employed, chymotryptic peptides showed maximum inhibition for angiotensin converting enzyme (ACE). The degree of hydrolysis and release kinetics of the peptides during chymotrypsin hydrolysis was compared with biological activity and the potent peptides fractions were identified. The crude fraction obtained after 110 min of hydrolysis shows multifunctional activities, like ACE inhibition, antioxidant activity, prolyl endopeptidase inhibitory activity and antimicrobial activities. This fraction was further purified by HPLC and sequenced by mass spectra. This fraction constituted peptides with molecular weights of 1,205, 1,718 Da respectively. The sequencing of peptides by MALDI-TOF MS/MS shows sequences QKALNEINQF and TKKTKLTEEEKNRL from α-_S2 casein.     

Molecular Modeling of the Ternary Complex of Rusticyanin-Cytochrome c4-Cytochrome Oxidase: An Insight to Possible H-Bond Mediated Recognition and Electron Transfer Reaction in T.ferrooxidans

Abstract

The metabolism of Thiobacillus ferrooxidans involves electron transfer from the Fe⁺² ions in the extracellular environment to the terminal oxygen in the bacterial cytoplasm through a series of periplasmic proteins like Rusticyanin (RCy), Cytochrome (Cyt c₄), and Cytochrome oxidase (CcO). The energy minimization and MD studies reveal the stabilization of the three redox proteins in their ternary complex through the direct and water mediated H-bonds and electrostatic interaction. The surface exposed polar residues of the three proteins, i.e., RCy (His 143, Thr 146, Lys 81, Glu 20), Cyt c₄ (Asp 5, 15, 52, Ser 14, Glu 61), and CcO (Asp 135, Glu 126, 140, 142, Thr 177) formed the intermolecular hydrogen bonds and stabilized the ternary complex. The oxygen (O^εl) of Glu 126, 140, and 142 on subunit II of the CcO interact to the exposed side-chain and O^b atoms of the Asp 52 of Cyt c₄ and Glu 20 and Leu 12 of RCy. The Asp 135 of subunit II also forms H-bond with the N^ε atom of Lys 81 of RCy. The O^εl of Glu 61 of Cyt c₄ is also H-bonded to O^γ atom of Thr 177 of CcO. Solvation followed by MD studies of the ternary protein complex revealed the presence of seven water molecules in the interfacial region of the interacting proteins. Three of the seven water molecules (W 79, W 437, and W 606) bridged the three proteins by forming the hydrogen bonded network (with the distances ～ 2.10–2.95 Å) between the Lys 81 (RCy), Glu 61 (Cyt c₄), and Asp 135 (CcO). Another water molecule W 603 was H-bonded to Tyr 122 (CcO) and interconnected the Lys 81 (RCy) and Asp 135 (CcO) through the water molecules W 606 and W 437. The other two water molecules (W 21 and W 455) bridged the RCy to Cyt c₄ through H-bonds, whereas the remaining W 76 interconnected the His 53 (Cytc₄) to Glu 126 (CcO) with distances ～ 2.95–3.0 Å.     

Formation mechanism of cross‐linking Maillard compounds in peptide–xylose systems

The formation mechanism of Maillard peptides was explored in Maillard reaction through diglycine/glutathione(GSH)/(Cys‐Glu‐Lys‐His‐Ile‐Met)–xlyose systems by heating at 120 °C for 30–120 min. Maximum fluorescence intensity of Maillard reaction products (MRPs) with an emission wavelength of 420~430 nm in all systems was observed, and the intensity values were proportional to the heating time. Taken diglycine/GSH–[¹³C₅]xylose systems as a control, it was proposed that the compounds with high m/z values of 379 and 616 have the high molecular weight (HMW) products formed by cross‐linking of peptides and sugar. In (Cys‐Glu‐Lys‐His‐Ile‐Met)–xylose system, the m/z value of HMW MRPs was not observed, which might be due to the weak signals of these products. According to the results of gel permeation chromatography, HMW MRPs were formed by Maillard reaction, especially in (Cys‐Glu‐Lys‐His‐Ile‐Met)–xylose system, the percentage of Maillard peptides reached 52.90%. It was concluded that Maillard peptides can be prepared through the cross‐linking of sugar and small peptides with a certain MW range. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.     

Insight into the binding of ACE-inhibitory peptides to angiotensin-converting enzyme: a molecular simulation

Angiotensin-I-converting enzyme (EC 3.4.15.1; ACE) plays an important physiological role in the regulation of blood pressure by converting angiotensin I to angiotensin II, a potent vasoconstrictor. Therefore, ACE inhibition has become a major target control for hypertension. Pipefish, or hailong, is an essential traditional Chinese medicine that is widely used in anti-fatigue and anti-cancer. A recent study has found two ACE-inhibitory peptides (TFPHGP and HWTTQR) purified from the seaweed pipefish by Alcalase enzymatic hydrolysis. Two peptides exhibited different ACE-inhibitory activities; however, the molecular mechanism involved is poorly understood. Further investigations are necessary to elucidate the relationship between the inhibition mechanism and the peptides. The current study is focused on investigating the interactions between each ACE-inhibitory peptide and ACE by performing molecular docking and molecular dynamics (MD) simulations. ACE protein remained stable throughout the simulations. Furthermore, ACE-TFPHGP complex showed lower binding energy as compared to ACE-HWTTQR complex, which is in good agreement with the experimental results. Glu384 and Glu411 of ACE are key residues upon the ACE-inhibitory peptides binding. Molecular basis generated by this attempt could provide valuable information towards designing new medicine for ACE inhibitor.     

Computer Modelling Studies on the Mechanism of Action of Ribonuclease T1

Abstract

The mechanism of action of ribonuclease (RNase) T₁ is still a matter of considerable debate as the results of x-ray, 2-D nmr and site-directed mutagenesis studies disagree regarding the role of the catalytically important residues. Hence computer modelling studies were carried out by energy minimisation of the complexes of RNase T₁ and some of its mutants (His40Ala, His40Lys, and Glu58Ala) with the substrate guanyl cytosine (GpC), and of native RNase T₁ with the reaction intermediate guanosine 2′, 3′-cyclic phosphate (G>p). The puckering of the guanosine ribose moiety in the minimum energy conformer of the RNase T₁ - GpC (substrate) complex was found to be O4′-endo and not C3′-endo as in the RNase T₁ - 3′-guanylic acid (inhibitor/product) complex. A possible scheme for the mechanism of action of RNase T₁ has been proposed on the basis of the arrangement of the catalytically important amino acid residues His40, Glu58, Arg77, and His92 around the guanosine ribose and the phosphate moiety in the RNase T₁ - GpC and RNase T₁ - G>p complexes. In this scheme, Glu58 serves as the general base group and His92 as the general acid group in the transphosphorylation step. His40 may be essential for stabilising the negatively charged phosphate moiety in the enzyme-transition state complex.     

Characterization of the particles of purified kappa-casein: trypsin as a probe of surface-accessible residues

-Casein as purified from bovine milk exhibits a rather unique disulfide bonding pattern as revealed by SDS–PAGE. The disulfide-bonded caseins present range from dimer to octamer and above and preparations contain about 10% monomer. All of these heterogeneous polymers, however, self-associate into nearly spherical particles with an average diameter of 13 nm at pH 8.0, as revealed by negatively stained transmission electron micrographs and dynamic light scattering. The weight-average molecular weight of the aggregates at pH 8.0, as judged by analytical ultracentrifugation, is 648,000. Trypsin digestion at pH 8.0 was used to probe the surface groups of the -casein A polymers. The reaction with trypsin was rapid and the peptides liberated were identified by separation with reverse-phase HPLC, amino acid analysis, and protein sequencing. The most rapidly released peptides (t _1/2 < 30 sec) were from cleavage at Arg 97 and Lys residues 111 and 112. These results suggest a surface orientation for these residues, and the data are in accord with earlier proposed 3D predictive models for -casein. It is speculated that Arg 97, together with adjacent His residues (98 and 100) and Lys residues 111 and 112, form two positively charged clusters on the surface of the otherwise negatively charged casein. These clusters bracket the neutral chymosin cleavage site (whose hydrolysis triggers a well-known digestive process) and so these clusters may facilitate docking of the substrate caseins with chymosin.     

Antihypertensive Peptides Derived from Soy Protein by Fermentation

Soy protein is widely used as a nitrogen source in infant and adult formulations, both in an intact and hydrolyzed form. Here, the objective was to screen for maximum proteolytic activity in different strains of lactobacillus and use it for fermentation of soy protein to obtain Angiotensin converting-I-enzyme (ACE I) inhibitory peptides for its use as a nutraceutical. Based on the proteolytic activity, Lactobacillus casei spp. pseudoplantarum was selected. The two ACE inhibitory peptide fractions F2 and F3 were isolated having IC₅₀ values of 17 ± 0.63 and 30 ± 0.13 μg/ml respectively. The N-terminal sequence of peptide (F2) was determined to be Leu-Ile-Val-Thr-Gln (LIVTQ). The peptide analogues of LIVTQ were synthesized to study the effect of individual residues on ACE enzyme. LIVTQ and LIVT peptides show inhibition against ACE enzyme having an IC₅₀ value of 0.087 and 0.110 μM respectively. Our results depict that glutamine (Q) and threonine (T) residues have an important role in ACE inhibition.     

The primary structure of the COOH-terminal half of cholera toxin subunit A1 containing the ADP-ribosylation site

The sequence of 96 amino acid residues from the COOH-terminus of the active subunit of cholera toxin, A₁, has been determined as PheAsnValAsnAspVal LeuGlyAlaTyrAlaProHisProAsxGluGlu GluValSerAlaLeuGlyGly IleProTyrSerGluIleTyrGlyTrpTyrArg ValHisPheGlyValLeuAsp GluGluLeuHisArgGlyTyrArgAspArgTyr TyrSerAsnLeuAspIleAla ProAlaAlaAspGlyTyrGlyLeuAlaGlyPhe ProProGluHisArgAlaTrp ArgGluGluProTrpIleHisHisAlaPro ProGlyCysGlyAsnAlaProArg(OH). This is the largest fragment obtained by BrCN cleavage of the subunit A₁ (M_r 23,000), and has previously been indicated to contain the active site for the adenylate cyclase-stimulating activity. Unequivocal identification of the COOH-terminal structure was achieved by separation and analysis of the terminal peptide after the specific chemical cleavage at the only cysteine residue in A₁ polypeptide. The site of self ADP-ribosylation in the A₁ subunit [C. Y. Lai, Q.-C. Xia, and P. T. Salotra (1983) Biochem. Biophys. Res. Commun.116, 341–348] has now been identified as Arg-50 of this peptide, 46 residues removed from the COOH-terminus. The cysteine that forms disulfide bridge to A₂ subunit in the holotoxin is at position 91.     

Constant pH molecular dynamics (CpHMD) and molecular docking studies of CquiOBP1 pH-induced ligand releasing mechanism

The odorant binding protein of Culex quinquefasciatus (CquiOBP1), expressed on the insect antenna, is crucial for the investigation of trapping baited with oviposition semi-chemicals and controlling mosquito populations. The acidic titratable residues pKa prediction and the ligand binding poses investigation in two systems (pH 7 and pH 5) are studied by constant pH molecular dynamics (CpHMD) and molecular docking methods. Research results reveal that the change of the protonation states would disrupt some important H-bonds, such as Asp 66-Asp 70, Glu 105-Asn 102, etc. The cleavage of these H-bonds leads to the movement of the relative position of hydrophobic tunnel, N- and C- termini loops and pH-sensing triad (His23-Tyr54-Val125) in acid solution. Ligand MOP has lower affinity and shows different binding poses to protein CquiOBP1 at pH 5. This ligand may be released from another tunnel between helices α3 and α4 in acidic environment. However, it would bind to the protein with high affinity in neutral environment. This work could provide more penetrating understanding of the pH-induced ligand-releasing mechanism.     

In vitro and in vivo ACE inhibitory of pistachio hydrolysates and in silico mechanism of identified peptide binding with ACE

The ACE inhibitory activity of pistachio (Pistacia vera L.) kernel's hydrolysates by gastrointestinal enzymes was studied. Results indicated that hydrolysate successively hydrolyzed by pepsin and trypsin, Pe–Tr–H, presented in vitro ACE inhibitory activity as IC₅₀ 0.87 ± 0.04 mg/ml. The Pe–Tr–H can in vivo decrease around 22 mmHg in systolic blood pressure (SBP) and 16 mmHg in the diastolic blood pressure (DBP) at 4 h after the oral administration, however the pistachio kernel powder can slightly lower SBP and DBP. The Pe–Tr–H with the highest activity was then separated by ultrafiltration membrane of 3 kDa, size exclusion chromatography on Sephadex G-15 and G-10 columns and reversed phase high-performance liquid chromatography (RP-HPLC) consecutively. A novel ACE inhibitory peptide, ACKEP, with the IC₅₀ value of 126 μM, was identified by MALDI–TOF/TOF system. ACKEP has the same C-terminal residue as Lisinopril and Enalapril, which plays a key role in binding with ACE. The binding mechanism was explored at a molecular basis by docking experiments, which revealed that seven residues from ACE active site (His383, His387, Glu384, Arg522, Asp358, Ala356 and Asn70) and two atoms of ACKEP (O5, H60) greatly contributed to the combinative stabilization.     

The molecular mechanisms of interactions between bioactive peptides and angiotensin-converting enzyme

The ability of milk protein derived Ile-Pro-Ala (IPA), Phe-Pro (FP) and Gly-Lys-Pro (GKP) peptides to inhibit angiotensin I-converting enzyme (ACE), a protein with an important role in blood-pressure regulation, were verified in vitro and in vivo. This work elucidates the modes and molecular mechanisms of the interaction of IPA, FP and GKP with ACE, including mechanisms that bind the peptides to the cofactor Zn²⁺. It was observed that the best docking poses obtained for IPA, FP and GKP were at the ACE catalytic site with very similar modes of interaction, including the interaction with Zn²⁺. The interactions, including H-bonds, hydrophobic, hydrophilic, and electrostatic interactions, as well as the interaction with Zn²⁺, were responsible for the binding between the bioactive peptides and ACE.