Identification of a cryptic functional apolipophorin-III domain within the Prominin-1 gene of Litopenaeus vannamei.

The Apolipophorin-III (apoLp-III) is reported as an essential protein element in lipids transport and incorporation in lepidopterans. Structurally, apoLp-III has an α-helix bundle structure composed of five α-helices. Interestingly, classic studies proposed a structural switch triggered by its interaction with lipids, where the α-helix bundle opens. Currently, the study of the apoLp-III has been limited to insects, with no homologs identified in other arthropods. By implementing a structure-based search with the Phyre2 algorithm surveying the shrimp Litopenaeus vannamei's transcriptome, we identified a putative apoLp-III in this farmed penaeid (LvApoLp-III). Unlike canonical apoLp-III, the LvApoLp-III was identified as an internal domain within the transmembrane protein Prominin-1. Structural modeling using the template-based Phyre2 and template-free AlphaFold algorithms rendered two distinct structural topologies: the α-helix bundle and a coiled-coil structure. Notably, the secondary structure composition on both models was alike, with differences in the orientation and distribution of the α-helices and hydrophobic moieties. Both models provide insights into the classical structural switch induced by lipids in apoLp-III. To corroborate structure/function inferences, we cloned the synthetic LvApoLp-III domain, overexpressed, and purified the recombinant protein. Circular dichroism measurements with the recombinant LvApoLp-III agreed with the structural models. In vitro liposome interaction demonstrated that the apoLp-III domain within the PROM1 of L.vannamei associated similarly to exchangeable apolipoproteins. Altogether, this work reports the presence of an apolipophorin-III domain in crustaceans for the first time and opens questions regarding its function and importance in lipid metabolism or the immune system.

Plant organellar MSH1 is a displacement loop specific endonuclease.

MSH1 is an organellar targeted protein that obstructs ectopic recombination and the accumulation of mutations in plant organellar genomes. MSH1 also modulates the epigenetic status of nuclear DNA, and its absence induces a variety of phenotypic responses. MSH1 is a member of the MutS family of DNA mismatch repair proteins but harbors an additional GIY-YIG nuclease domain that distinguishes it from the rest of this family. How MSH1 hampers recombination and promotes fidelity in organellar DNA inheritance is unknown. Here, we elucidate its enzymatic activities by recombinantly expressing and purifying full-length MSH1 from Arabidopsis thaliana (AtMSH1). AtMSH1 is a metalloenzyme that shows a strong binding affinity for displacement loops (D-loops). The DNA binding abilities of AtMSH1 reside in its MutS domain and not in its GIY-YIG domain, which is the ancillary nickase of AtMSH1. In the presence of divalent metal ions, AtMSH1 selectively executes multiple incisions at D-loops, but not other DNA structures including Holliday junctions or dsDNA, regardless of the presence or absence of mismatches. The selectivity of AtMSH1 to dismantle D-loops supports the role of this enzyme in preventing recombination between short repeats. Our results suggest that plant organelles have evolved novel DNA repair routes centered around the anti-recombinogenic activity of MSH1.

TtCel7A: A Native Thermophilic Bifunctional Cellulose/Xylanase Exogluclanase from the Thermophilic Biomass-Degrading Fungus Thielavia terrestris Co3Bag1, and Its Application in Enzymatic Hydrolysis of Agroindustrial Derivatives.

The biomass-degrading thermophilic ascomycete fungus Thielavia terrestris Co3Bag1 produces TtCel7A, a native bifunctional cellulase/xylanase GH7 family. The purified TtCel7A, with an estimated molecular weight of 71 kDa, was biochemically characterized. TtCel7A displayed an optimal pH of 5.5 for both activities and an optimal temperature of 60 and 50 °C for cellulolytic and xylanolytic activities, respectively. The half-lives determined for cellulase activity were 140, 106, and 41 min at 50, 60, and 70 °C, respectively, whereas the half-lives observed for xylanase activity were 24, 10, and 1.4 h at 50, 60, and 70 °C, respectively. The KM and Vmax values were 3.12 mg/mL and 50 U/mg for cellulase activity and 0.17 mg/mL and 42.75 U/mg for xylanase activity. Circular dichroism analysis suggests changes in the secondary structure of TtCel7A in the presence of CMC as the substrate, whereas no modifications were observed with beechwood xylan. TtCel7A displayed the excellent capability to hydrolyze CMC, beechwood xylan, and complex substrates such as oat bran, wheat bran, and sugarcane bagasse, with glucose and cellobiose being the main products released; also, slightly less endo cellulase and xylanase activities were observed. Thus, suggesting TtCel7A has an exo- and endomode of action. Based on the characteristics of the enzyme, it might be considered a good candidate for industrial applications.

Atomistic mechanism of leptin and leptin-receptor association.

The leptin-leptin receptor complex is at the very core of energy homeostasis and immune system regulation, among many other functions. In this work, we built homology models of leptin and the leptin binding domain (LBD) of the receptor from humans and mice. Docking analyses were used to obtain the coordinates of the native leptin-LBD complexes and a mixed heterodimer formed by human leptin and mouse LBD. Molecular dynamics (MD) simulations were performed using all models (monomers and heterodimers) as initial coordinates and the GROMACS program. The overall structural and dynamical behaviors are similar for the three complexes. Upon MD simulations, several new interactions appear. In particular, hydrophobic interactions, with more than 90% persistence, seem to be the most relevant for the stability of the dimers, as well as the pair formed by Asp85Lep and Arg468LBD. This in silico analysis provides structural and dynamical information, at the atomistic level, about the mechanism of leptin-LBD complex formation and leptin receptor activation. This knowledge might be used in the rational drug design of therapeutics to modulate leptin signaling.Communicated by Ramaswamy H. Sarma.

Anti-apoptotic Bcl-2 protein in apo and holo conformation anchored to the membrane: comparative molecular dynamics simulations.

The interaction between the anti-apoptotic Bcl-2 protein and its antagonist Bax is essential to the regulation of the mitochondrial pathway of apoptosis. For this work, we built models by homology of Bcl-2 full-sequence length in monomeric form (apo-Bcl-2) and in complex with the BH3 domain of Bax (holo-Bcl-2). The Bcl-2 protein was analyzed with its transmembrane domain anchored to a lipidic bilayer of DPPC, imitating physiological conditions. We performed molecular dynamics (MD) simulations using the GROMACS program. Conformational changes showed that the flexible loop domain (FLD) tends to fold on itself and move towards the main core. Furthermore, the BH3 peptide of pro-apoptotic protein Bax, showed an allosteric stabilizing effect on FLD upon being bound to the hydrophobic cleft of the anti-apoptotic protein Bcl-2, causing a reduction in its structural flexibility. However, FLD is distal from the main core of Bcl-2. Principal component analysis (PCA) showed a weak correlation between FLD residues and BH3 peptide from Bax. Upon MD simulations, several new contacts appeared between FLD and some α-helices of the core of Bcl-2, which contribute to maintaining the stability of Bcl-2. This knowledge sheds light on the behavior of Bcl-2 in the cell's native environment.Communicated by Ramaswamy H. Sarma.

Expression, purification, and refolding of the recombinant extracellular domain ß1-subunit of the dog Na+/K+-ATPase of the epithelial cells.

The ß1-subunit of the Na+/K+-ATPase is a cell membrane protein, beyond its classic functions, it is also a cell adhesion molecule. ß1-subunits on the lateral membrane of dog kidney epithelial cells trans-interact with ß1-subunits from another neighboring cells. The ß-ß interaction is essential for the formation and stabilization of intercellular junctions. Previous studies on site-directed mutagenesis and in silico revealed that the interaction interface involves residues 198-207 and 221-229. However, it is necessary to report the interaction interface at the structural level experimentally. Here, we describe the successful cloning, overexpression in E. coli, and purification of the extracellular domain of the ß1-subunit from inclusion bodies. Experimental characterization by size exclusion chromatography and DLS indicated similar hydrodynamic properties of the protein refolded. Structural analysis by circular dichroism and Raman spectroscopy revealed the secondary structures in the folded protein of type ß-sheet, α-helix, random coil, and turn. We also performed ß1-ß1 interaction assays with the recombinant protein, showing dimers' formation (6xHisß1-ß1). Given our results, the recombinant extracellular domain of the ß1-subunit is highly similar to the native protein, therefore the current work in our laboratory aims to characterize at the atomic level the interaction interface between EDß1.

Foaming and Structural Studies on the Acidic Subunit of Amaranth 11S Globulin Modified with Antihypertensive Peptides as a Function of pH and Ionic Strength.

Some studies aimed at revealing the relationship between protein structure and their functional properties. However, the majority of these reports have been carried out using protein isolates. There are limited reports on the possible relationship between the functional properties and the structure of a purified protein. In this work the amaranth 11S globulin acidic subunit (AAC) and five mutations of the same protein that were modified in their variable regions with antihypertensive peptides (VYVYVYVY and RIPP), were analyzed at two ionic strength (2.9 and 17.6 g/L NaCl) and pH (3.0-7.0). Results revealed better solubility for the proteins mutated at the terminal ends (AACM.1 and AACM.4) and lower solubility for the protein inserted with RIPP peptide. Spectroscopy studies revealed an increase of ß-sheet structure at high salt concentration for all proteins. It was also observed that salt concentration acted as a modulator, which allowed a better foam features for all modified proteins limiting movement of side chains and reducing red-shifted displacement of λmax. All proteins showed foam capacity ranging from 76 to 93% although foam stability was twofold better for modified proteins than for AAC at high salt concentration. This study allowed better understanding about the structural changes that influence the foaming properties of engineered proteins.

Improved Stability of Human CGI-58 Induced by Phosphomimetic S237E Mutation.

In lipolysis, the activating function of CGI-58 is regulated by its interaction with perilipin 1 (PLIN1) localized on the lipid droplet (LD), and its release is controlled by phosphorylation. Once lipolysis is stimulated by catecholamines, protein kinase A (PKA)-mediated phosphorylation enables the dissociation of the CGI-58/PLIN1 complex, thereby recruiting adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) to initiate fatty acid release. It has been shown that mouse CGI-58 mutant S239E, which mimics the phosphorylation of this residue, is able to dissociate from the CGI-58/PLIN1 complex and activate ATGL. Here, we analyze the stabilizing effect on human CGI-58 of a triple tryptophan to alanine mutant (3WA) on the LD-binding motif, as well as a quadruple mutant in which the phosphomimetic S237E substitution was introduced to the 3WA construct (3WA/S237E). We found that tryptophan residues promote wild-type (WT) protein aggregation in solution since their substitution for alanine residues favors the presence of the monomer. Our experimental data showed increased thermal stability and solubility of 3WA/S237E protein compared to the 3WA mutant. Moreover, the 3WA/S237E protein showed proper folding and a functional binding site for oleoyl-CoA. The analysis of a bioinformatic three-dimensional (3D) model suggests an intramolecular interaction between the phosphomimetic glutamic acid and a residue of the α/ß hydrolase core. This could explain the increased solubility and stability observed in the 3WA/S237E mutant and evidences the possible role of serine 237 phosphorylation.

Synergistic effect of compounds directed to triosephosphate isomerase, a combination to develop drug against trichomoniasis.

The development of new drugs is continuous in the world; currently, saving resources (both economic ones and time) and preventing secondary effects have become a necessity for drug developers. Trichomoniasis is the most common nonviral sexually transmitted infection affecting more than 270 million people around the world. In our research group, we focussed on developing a selective and more effective drug against Trichomonas vaginalis, and we previously reported on a compound, called A4, which had a trichomonacidal effect. Later, we determined another compound, called D4, which also had a trichomonacidal effect together with favorable toxicity results. Both A4 and D4 are directed at the enzyme triosephosphate isomerase. Thus, we made combinations between the two compounds, in which we determined a synergistic effect against T. vaginalis, determining the IC50 and the toxicity of the best relationship to obtain the trichomonacidal effect. With these results, we can propose a combination of compounds that represents a promising alternative for the development of a new therapeutic strategy against trichomoniasis.

The novel EhHSTF7 transcription factor displays an oligomer state and recognizes a heat shock element in the Entamoeba histolytica parasite.

The heat shock response is a conserved mechanism that allows cells to respond and survive stress damage and is transcriptionally regulated by the heat shock factors and heat shock elements. The P-glycoprotein confer the multidrug resistance phenotype; Entamoeba histolytica has the largest multidrug resistance gene family described so far; one of these genes, the EhPgp5 gene, has an emetine-inducible expression. A functional heat shock element was localized in the EhPgp5 gene promoter, indicating transcriptional regulation by heat shock factors. In this work, we determined the oligomer state of EhHSTF7 and the recognition of the heat shock element of the EhPgp5 gene. The EhHSTF7 recombinant protein was obtained as monomer and oligomer. In silico molecular docking predicts protein-DNA binding between EhHSTF7 and 5'-GAA-3' complementary bases. The rEhHSTF7 protein specifically binds to the heat shock element of the EhPgp5 gene in gel shift assays. The competition assays with heat shock element mutants indicate that 5'-GAA-3' complementary bases are necessary for the rEhHSTF7 binding. Finally, the siRNA-mediated knockdown of Ehhstf7 expression causes downregulation of EhPgp5 expression, suggesting that EhHSTF7 is likely to play a key role in the E. histolytica multidrug resistance. This is the first report of a transcription factor that recognizes a heat shock element from a gene involved in drug resistance in parasites. However, further analysis needs to demonstrate the biological relevance of the EhHSTF7 and the rest of the heat shock factors of E. histolytica, to understand the underlying regulation of transcriptional control in response to stress.

Differentially expressed proteins in platelets derived from patients with hypertension.

Hypertension (HTN) causes end-organ damage and is a major cause of morbidity and mortality globally. Recent studies suggested blood cells participate in the maintenance of HTN. Platelets-anucleated cell fragments derived from megakaryocytes-exert diverse functions, including their well-characterized role in the formation of hemostatic clots. However, platelets from patients with HTN exhibit altered membrane lipid and protein compositions that impact platelet function and lead to formation of aggregates and vascular obstructions. Here, for the first time, we have identified, by proteomic analyses, the most relevant 11 proteins that show the greatest difference in their expression in platelets derived from patients with HTN, in comparison with those from normotensive individuals. These proteins are involved in cytoskeletal organization and the coagulation cascade that contributes to platelet activation, release of granule contents, and aggregation, which culminate in thrombus formation. These results have important implications in our understanding of the molecular mechanisms associated with the development of HTN, and in consequence, the development of new strategies to counteract the cardiovascular disorders associated with constitutive activation of platelets in HTN.

Triosephosphate isomerase as a therapeutic target against trichomoniasis.

Trichomoniasis is the most common non-viral sexually transmitted infection, caused by the protozoan parasite Trichomonas vaginalis, affecting millions of people worldwide. The main treatment against trichomoniasis is metronidazole and other nitroimidazole derivatives, but up to twenty percent of clinical cases of trichomoniasis are resistant to these drugs. In this study, we used high-performance virtual screening to search for molecules that specifically bind to the protein, triosephosphate isomerase from T. vaginalis (TvTIM). By in silico molecular docking analysis, we selected six compounds from a chemical library of almost 500,000 compounds. While none of the six inhibited the enzymatic activity of recombinant triosephosphate isomerase isoforms, one compound (A4; 3,3'-{[4-(4-morpholinyl)phenyl]methylene}bis(4- hydroxy-2H-chromen-2-one) altered their fluorescence emission spectra, suggesting that this chemical might interfere in an important non-glycolytic function of TvTIM. In vitro assays demonstrate that A4 is not cytotoxic but does have trichomonacidal impact on T. vaginalis cultures. With these results, we propose this compound as a potential drug with a new therapeutic target against Trichomonas vaginalis.

Identifying compounds that prevent the binding of the SARS-CoV-2 S-protein to ACE2.

We investigated compounds selected by molecular docking to identify a specific treatment for COVID-19 that decreases the interaction between angiotensin-converting enzyme 2 (ACE2) and the receptor-binding domain (RBD) of SARS-CoV-2. Five compounds that interact with ACE2 amino acids Gln24, Asp30, His34, Tyr41, Gln42, Met82, Lys353, and Arg357 were evaluated using specific binding assays for their effects on the interaction between ACE2 with RBD. The compound labeled ED demonstrated favorable ACE2-binding, with an IC50 of 31.95 µM. ED cytotoxicity, evaluated using PC3 cells in an MTT assay, was consistent with the low theoretical toxicity previously reported. We propose that ED mainly interacts with His34, Glu37, and Lys353 in ACE2 and that it has an inhibitory effect on the interaction of ACE2 with the RBD of the S-protein. We recommend further investigation to develop ED into a potential drug or adjuvant in COVID-19 treatment.

Effect of the single mutation N9Y on the catalytical properties of xylanase Xyn11A from Cellulomonas uda: a biochemical and molecular dynamic simulation analysis.

Cellulomonas uda produces Xyn11A, moderately thermostable xylanase, with optimal activity at 50 °C and pH 6.5. An improvement in the biochemical properties of Xyn11A was achieved by site-directed mutagenesis approach. Wild-type xylanase, Xyn11A-WT, and its mutant Xyn11A-N9Y were expressed in Escherichia coli, and then both enzymes were purified and characterized. Xyn11A-N9Y displayed optimal activity at 60 °C and pH 7.5, an upward shift of 10 °C in the optimum temperature and an upward shift of 1 unit in optimum pH; also, it manifested an 11-fold increase in thermal stability at 60 °C, compared to that displayed by Xyn11A-WT. Molecular dynamics simulations of Xyn11A-WT and Xyn11A-N9Y suggest that the substitution N9Y leads to an array of secondary structure changes at the N-terminal end and an increase in the number of hydrogen bonds in Xyn11A-N9Y. Based on the significant improvements, Xyn11A-N9Y may be considered as a candidate for several biotechnological applications.

Structure of an open conformation of T7 DNA polymerase reveals novel structural features regulating primer-template stabilization at the polymerization active site.

The crystal structure of full-length T7 DNA polymerase in complex with its processivity factor thioredoxin and double-stranded DNA in the polymerization active site exhibits two novel structural motifs in family-A DNA polymerases: an extended ß-hairpin at the fingers subdomain, that interacts with the DNA template strand downstream the primer-terminus, and a helix-loop-helix motif (insertion1) located between residues 102 to 122 in the exonuclease domain. The extended ß-hairpin is involved in nucleotide incorporation on substrates with 5'-overhangs longer than 2 nt, suggesting a role in stabilizing the template strand into the polymerization domain. Our biochemical data reveal that insertion1 of the exonuclease domain makes stabilizing interactions that facilitate proofreading by shuttling the primer strand into the exonuclease active site. Overall, our studies evidence conservation of the 3'-5' exonuclease domain fold between family-A DNA polymerases and highlight the modular architecture of T7 DNA polymerase. Our data suggest that the intercalating ß-hairpin guides the template-strand into the polymerization active site after the T7 primase-helicase unwinds the DNA double helix ameliorating the formation of secondary structures and decreasing the appearance of indels.

Esters of quinoxaline-7-carboxylate-1,4-di-N-oxide as Trichomonas vaginalis triosephosphate isomerase inhibitors.

Trichomoniasis is a public health problem worldwide. Trichomoniasis treatment consists of the use of nitroimidazole derivatives; however, therapeutic ineffectiveness occurs in 5 to 20 % of the cases. Therefore, it is essential to propose new pharmacological agents against this disease. In this work, esters of quinoxaline-7-carboxylate-1,4-di-N-oxide (EQX-NO) were evaluated in in vitro assays as novel trichomonicidal agents. Additionally, an in vitro enzyme assay and molecular docking analysis against triosephosphate isomerase of Trichomonas vaginalis to confirm their mechanism of action were performed. Ethyl (compound 12) and n-propyl (compound 37) esters of quinoxaline-7-carboxy-late-1,4-di-N-oxide derivatives showed trichomonicidal activity comparable to nitazoxanide, whereas five methyl (compounds 5, 15, 19, 20 and 22), four isopropyl (compounds 28, 29, 30 and 34), three ethyl (compounds 4, 13 and 23) and one npropyl (compound 35) ester derivatives displayed activity comparable to albendazole. Compounds 6 and 20 decreased 100 % of the enzyme activity of recombinant protein triosephosphate isomerase.

Amoebicidal effect of 5,5'-[(4-nitrophenyl)methylene]bis-6-hydroxy-2-mercapto-3-methyl-4(3H)-pyrimidinone), a new drug against Entamoeba histolytica.

Entamoeba histolytica is a cosmopolitan protozoan parasite that can produce infections in the intestine and some organs (liver, lungs, and brain), with worldwide prevalence. There are treatments against E. histolytica (antiparasitics), but as the drugs used in these treatments have presented some type of resistance and/or side effects, there are cases with complications of this disease. Therefore, it is necessary to develop new drugs aimed at a specific therapeutic target against this parasite. Here, we used the compound 5,5'-[(4-nitrophenyl)methylene]bis(6-hydroxy-2-mercapto-3-methyl-4(3H)-pyrimidinone) in the patenting process (called D4). D4 has a reported specific use against a glycolytic enzyme, the triosephosphate isomerase of Trichomonas vaginalis (TvTIM). We determined that D4 has an amoebicidal effect in in vitro cultures, with an IC50 value of 18.5 µM, and we proposed a specific site of interaction (Lys77, His110, Gln115, and Glu118) in the triosephosphate isomerase of E. histolytica (EhTIM). Furthermore, compound D4 has favorable experimental and theoretical toxicity results. Therefore, D4 should be further investigated as a potential drug against E. histolytica.

Bioinformatic Analysis and Biophysical Characterization Reveal Structural Disorder in G0S2 Protein.

G0S2 is a small protein of 103 residues in length that is involved in multiple cellular processes. To date, several reports have shown that G0S2 functions by making direct protein-protein interactions with key proteins. In lipolysis, G0S2 specifically interacts with adipose triglyceride lipase, inhibiting its activity and resulting in lipolysis being downregulated. In a similar way, G0S2 also participates in the regulation of apoptosis, cell proliferation, and oxidative phosphorylation; however, information regarding G0S2 structural and biophysical properties is limited. In this work, we conducted a comparative structural analysis of human and mouse G0S2 proteins. Bioinformatics suggests the presence of a disordered C-terminal region in human G0S2. Experimental characterization by size-exclusion chromatography and dynamic light scattering showed that human and mouse G0S2 have different hydrodynamic properties. In comparison to the mouse G0S2, which behaves similar to a globular protein, the human G0S2 shows an elongated conformation, most likely by displaying a disordered C-terminal region. Further analysis of hydrodynamic properties under denaturing conditions suggests the presence of a structural element in the mouse protein that undergoes an order to disorder transition at low urea concentration. Structural analysis by circular dichroism revealed that in native conditions, both proteins are mainly unstructured, showing the presence of beta sheet structures. Further analysis of CD data suggests that both proteins belong to the premolten globule family of intrinsically disordered proteins. We suggest that the intrinsic disorder observed in the G0S2 protein may facilitate its interaction with multiple partners in the regulation of cellular metabolism.

Solution structure of the inhibitor of cysteine proteases 1 from Entamoeba histolytica reveals a possible auto regulatory mechanism.

The genome of Entamoeba histolytica encodes approximately 50 Cysteine Proteases (CPs) whose activity is regulated by two Inhibitors of Cysteine Proteases (ICPs), EhICP1 and EhICP2. The main difference between both EhICPs is the acquisition of a 17 N-terminal targeting signal in EhICP2 and three exposed cysteine residues in EhICP1. The three exposed cysteines in EhICP1 potentiate the formation of cross-linking species that drive heterogeneity. Here we solved the NMR structure of EhICP1 using a mutant protein without accessible cysteines. Our structural data shows that EhICP1 adopts an immunoglobulin fold composed of seven ß-strands, and three solvent exposed loops that resemble the structures of EhICP2 and chagasin. EhICP1 and EhICP2 are able to inhibit the archetypical cysteine protease papain by intercalating their BC loops into the protease active site independently of the character of the residue (serine or threonine) responsible to interact with the active site of papain. EhICP1 and EhICP2 present signals of functional divergence as they clustered in different clades. Two of the three exposed cysteines in EhICP1 are located at the DE loop that intercalates into the CP substrate-binding cleft. We propose that the solvent exposed cysteines of EhICP1 play a role in regulating its inhibitory activity and that in oxidative conditions, the cysteines of EhICP1 react to form intra and intermolecular disulfide bonds that render an inactive inhibitor. EhICP2 is not subject to redox regulation, as this inhibitor does not contain a single cysteine residue. This proposed redox regulation may be related to the differential cellular localization between EhICP1 and EhICP2.

Potential inhibitors of the interaction between ACE2 and SARS-CoV-2 (RBD), to develop a drug.

AIMS: The COVID-19 disease caused by the SARS-CoV-2 has become a pandemic and there are no effective treatments that reduce the contagion. It is urgent to propose new treatment options, which are more effective in the interaction between viruses and cells. In this study was to develop a search for new pharmacological compounds against the angiotensin-converting enzyme 2 (ACE2), to inhibit the interaction with SARS-CoV-2. MATERIALS AND METHODS: Docking, virtual screening using almost 500,000 compounds directed to interact in the region between the residues (Gln24, Asp30, His34, Tyr41, Gln42, Met82, Lys353, and Arg357) in ACE2. The average of ΔGbinding, the standard deviation value and the theoretical toxicity from compounds were analyzed. KEY FINDINGS: 20 best compounds directed to interact in ACE2 with a high probability to be safe in humans, validated by web servers of prediction of ADME and toxicity (ProTox-II and PreADMET), to difficult the interaction between ACE2 and region binding domain (RBD) of SARS-CoV-2. SIGNIFICANCE: In this study, 20 compounds were determined by docking focused on the region of interaction between ACE2 and RBD of SARS-CoV-2 was carried out. The compounds are publicly available to validate the effect in in vitro tests.