X-ray Structure Determination Research Articles

Synthesis of bismuthanyl-substituted monomeric triel hydrides.

The syntheses and characterizations of the first bismuthanylborane monomers stabilized only by a donor in D·BH2Bi(SiMe3)2 (D = DMAP 1a, IDipp 1b, IMe41c; DMAP = 4-dimethylaminopyridine, IDipp = 1,3-bis(2,6-diisopropylphenyl)-imidazolin-2-ylidene, IMe4 = 1,3,4,5-tetramethylimidazol-2-ylidene) are presented. All compounds were synthesized by salt metathesis reactions between D·BH2I and KBi(SiMe3)2(THF)0.3 and represent some of the extremely rare compounds featuring a 2c-2e B-Bi bond in a molecular compound. The products display high sensitivity towards air and light and slowly decompose in solution even at -80 °C. By the reaction of IDipp·GaH2(SO3CF3) with KBi(SiMe3)2(THF)0.3, the synthesis of the first bismuthanylgallane IDipp·GaH2Bi(SiMe3)2 (2) stabilized only by a 2-electron donor was possible, as evident from single crystal X-ray structure determination, NMR spectroscopy and mass spectrometry. Computational studies shed light on the stability of the products and the electronic nature of the compounds.

Structural consequences in differently substituted haloarylcarboxylates and non-covalent interactions on crystal packing and biological efficacy of copper(II) complexes with N,N,N′,N′-tetramethylethylenediamine N-donor ligand

In this work, we reported four new copper(II) halo-benzoate complexes (2-chloro-4-nitrobenzoate (2-Cl-4-NO2-BZ), 3,5-DICBZ, 2-chloro-5-nitrobenzoate (2-Cl-5-NO2-BZ), 3,5-difluoro benzoate (3,5-DIFBZ), and 3,5-dichlorobenzene (3,5-DICBZ) respectively in complexes 1–4) with N,N,N’,N’- tetramethylethylene diamine (temed) N-donor ligand in order to study the interesting coordination features and biological evaluation of such complexes under ambient reaction conditions. All the complexes 1–4 were characterized by elemental analyses, and spectroscopic techniques, including FT-IR, UV–vis, and EPR etc. Single crystal X-ray structure determination (SCXRD) of complexes revealed the distorted octahedral geometry in all complexes. Notably, the halobenzoate ligands exhibited a bidentate chelation mode in complexes 1 and 2, while they showed a monodentate mode in complex 3. In complex 4, the carboxylate ligand exhibited both monodentate as well as bidentate coordination. Furthermore, detailed packing analyses of complexes 1–4 highlighted the significance of halogen bonding in lattice stabilization besides hydrogen bonding and π∙∙∙π interactions as evidenced by Hirshfeld surface analyses and crystal analysis. Moreover, packing analyses displayed that complex 1 exhibited a layered or wave-like arrangement, while complex 2 featured a helical arrangement supported by a supramolecular halogen-bonded network guided by Cl∙∙∙O interactions. Complex 3 displayed a zig-zag layered arrangement with a stacking topology, and Complex 4 showcased a ribbon-like arrangement stabilized by F∙∙∙F and Cl∙∙∙Cl halogen bonding interactions. Furthermore, sigma hole on halogen atoms has also been examined (which play an important role in halogen bonding) by electrostatic-potential isosurfaces. In order to exploit the biological efficacy of complexes 1–4 owing to the inherent biological activity of copper metal, molecular docking analyses against gram +ve bacteria i.e. S. epidermidis (PDB ID; 8DO6), B. subtilis (1QD9), as well as two gram -ve bacteria, namely, P. aeruginosa (5WZE) and S. dysenteriae (1DM0), have also been carried out. Interestingly, values of docking scores and inhibition constant of all complexes 1–4 (with a maximum binding score (inhibition constant) of -9.8 kcal/mol (0.063 µMol) in 1 against 8DO6 and 1DM0) revealed higher biological efficacy than that of standard drugs.

Synthesis, structure, preliminary antimicrobial and antimalarial studies of 1,1′-(piperazine-1,4-diyl)bis(2-phenylethan-1-one) and its lanthanide, Ce(III), Pr(III), and Nd(III) complexes

A dione ligand, 1,1′-(piperazine-1,4-diyl)bis(2-phenylethan-1-one) derived from the reaction of phenylacetylchloride with piperazine and the lanthanides, Ce(III), Nd(III), and Pr(III) complexes of general formula [LnL(NO3)2]NO3 were synthesized. The compounds were characterized by conventional NMR, Mass Infra-Red and UV–vis spectroscopic techniques, melting point, elemental analysis, conductivity, thermogravimetric analysis, magnetic susceptibility, and single X-ray structure determination of the ligand. In vitro, antimicrobial studies indicated that the ligand is active against Staphylococcus aureus, Bacillus Sublitis, Salmonella typhi, and Streptococcus pneumonae. The Ce(III) and Pr(III) complexes were active against all the microorganisms except A. Niger and Pseudomonas aeruginosa whereas the Ce(III) complex showed activity against Staphylococcus aureus, Bacillus Sublitis, and Streptococcus pneumonae. The In vivo anti-plasmodial assay of infected rats indicated that the ligand and Nd(III), Pr(III) complexes have high antiplasmodial potency in comparison to the antimalarial medicine Artesunate. In corroboration with findings on the enhanced antimicrobial properties of the complexes, binding energies from molecular docking correlated strongly with the inhibition zone diameter for Staphylococcus aureus and density functional theoretical studies suggest the presence of the metals account for the improved properties in the following order: Nd(III) > Pr(III) > Ce(III).

Structural and biochemical analysis of the PARP1-homology region of PARP4/vault PARP.

PARP4 is an ADP-ribosyltransferase that resides within the vault ribonucleoprotein organelle. Our knowledge of PARP4 structure and biochemistry is limited relative to other PARPs. PARP4 shares a region of homology with PARP1, an ADP-ribosyltransferase that produces poly(ADP-ribose) from NAD+ in response to binding DNA breaks. The PARP1-homology region of PARP4 includes a BRCT fold, a WGR domain, and the catalytic (CAT) domain. Here, we have determined X-ray structures of the PARP4 catalytic domain and performed biochemical analysis that together indicate an active site that is open to NAD+ interaction, in contrast to the closed conformation of the PARP1 catalytic domain that blocks access to substrate NAD+. We have also determined crystal structures of the minimal ADP-ribosyltransferase fold of PARP4 that illustrate active site alterations that restrict PARP4 to mono(ADP-ribose) rather than poly(ADP-ribose) modifications. We demonstrate that PARP4 interacts with vault RNA, and that the BRCT is primarily responsible for the interaction. However, the interaction does not lead to stimulation of mono(ADP-ribosylation) activity. The BRCT-WGR-CAT of PARP4 has lower activity than the CAT alone, suggesting that the BRCT and WGR domains regulate catalytic output. Our study provides first insights into PARP4 structure and regulation and expands understanding of PARP structural biochemistry.

Ethoxylate Polymer-Based 96-Well Screen for Protein Crystallization

Crystallization is the limiting step in X-ray structure determination of biological macromolecules. As crystallization experiments can be largely automatized, the diversity of precipitant solutions is often the determinant factor to obtain crystals of high quality. Here, we introduce a 96-well screening kit of crystallization conditions, centered on three ethoxylate-based organic polymers as precipitants and various additional compounds to promote crystal formation. This crystallization screen was tested on various non-standard proteins from bacteria and archaea. Structure determination succeeded for seven out of thirteen targets based on crystals that frequently diffracted to a higher resolution than those obtained with commercially available screening kits. Crystallization hits were rarely similar among the three ethoxylate-based organic polymers and, in comparison, with already available crystallization screens. Hence, the presented crystallization screen is an efficient tool to complement other screens and increase the likelihood of growing crystals suitable for X-ray structure determination.

X-ray structures of Enterobacter cloacae allose-binding protein in complexes with monosaccharides demonstrate its unique recognition mechanism for high affinity to allose

d-Allose is an aldohexose of the C3-epimer of d-glucose, existing in very small amounts in nature, called a rare sugar. The operon responsible for d-allose metabolism, the allose operon, was found in several bacteria, which consists of seven genes: alsR, alsB, alsA, alsC, alsE, alsK, and rpiB. To understand the biological implication of the allose operon utilizing a rare sugar of d-allose as a carbon source, it is important to clarify whether the allose operon functions specifically for d-allose or also functions for other ligands. It was proposed that the allose operon can function for d-ribose, which is essential as a component of nucleotides and abundant in nature. Allose-binding protein, AlsB, coded in the allose operon, is thought to capture a ligand outside the cell, and is expected to show high affinity for the specific ligand. X-ray structure determinations of Enterobacter cloacae AlsB (EtcAlsB) in ligand-free form, and in complexes with d-allose, d-ribose, and d-allulose, and measurements of the thermal parameters of the complex formation using an isothermal titration calorimeter were performed. The results demonstrated that EtcAlsB has a unique recognition mechanism for high affinity to d-allose by changing its conformation from an open to a closed form depending on d-allose-binding, and that the binding of d-ribose to EtcAlsB could not induce a completely closed form but an intermediate form, explaining the low affinity for d-ribose.

The synthesis of novel lanthanum hydroxyborate at extreme conditions.

The novel structure of lanthanum hydroxyborate La2B2O5(OH)2 was synthesized by the reaction of partially hydrolyzed lanthanum and boron oxide in a diamond anvil cell under high-pressure/high-temperature (HPHT) conditions of 30GPa and ∼2,400K. The single-crystal X-ray structure determination of the lanthanum hydroxyborate revealed: , a = 6.555(2)Å, c = 17.485(8)Å, Z = 6, R1 = 0.056. The three-dimensional structure consists of discrete planar BO3 groups and three crystallographically different La ions: one is surrounded by 9, one by 10, and one by 12 oxygen anions. The band gap was estimated using ab initio calculations to be 4.64eV at ambient pressure and 5.26eV at 30GPa. The current work describes the novel HPHT lanthanum hydroxyborate with potential application as a deep-ultraviolet birefringent material.

Cocrystal of Codeine and Cyclopentobarbital

The two-component compound formed by codeine and cyclopentobarbital was produced using grinding techniques and through evaporation from alcoholic solutions. The cocrystal nature of this phase was established unequivocally through single crystal X-ray structure determination. The asymmetric unit contains one formula unit. In the cyclopentobarbital molecule, the cyclopentenyl ring is disordered over two positions related by a rotation of approximately 180° about its C—C bond to the pyrimidine ring. The two NH groups of the cyclopentobarbital molecule form N—H⋯N and N—H⋯O bonds to piperidine and hydroxyl groups, respectively, belonging to different codeine molecules. In addition, the hydroxyl and methoxy groups of neighboring codeine molecules are linked by O—H⋯O interactions, resulting in a H-bonded framework structure of codeine and cyclopentobarbital molecules. The cocrystal was also characterized using thermal analysis, X-ray powder diffraction and IR spectroscopy.

Novel hybrid motifs of 4-nitroimidazole-piperazinyl tagged 1,2,3-triazoles: Synthesis, crystal structure, anticancer evaluations, and molecular docking study

4-((4-(1-benzyl-2-methyl-4-nitro-1H-imidazole-5-yl)piperazine-1-yl)methyl)-1-substituted-1H-1,2,3-triazole motifs are designed and synthesized via click chemistry. The reaction of 1-(N1-benzyl- 2-methyl-4-nitro-1H-imidazole- 5-yl)-4-(prop-2-yn-1-yl) piperazine 5 as new scaffold with diverse primary azides to selectively produce 1,4-disubstituted-1,2,3-triazoles 9a–k, 10a–c and 11a–q. Physicochemical methods: when 1H NMR, 13C NMR, and HRMS are utilized to fully characterize all synthesized compounds. X-ray structural determination and analysis for compound 9a is also performed. The newly designed chromophores are assessed for their anti-proliferative potency against three selected human cancer cell lines (MCF-7, HepG2, and PC3), and one normal cell line (Dermal/Fibroblast). Compounds 9g and 9k have shown potent activities against the MCF-7 cell line with IC50 values of (2.00 ± 0.03 μM) and (5.00 ± 0.01 μM) respectively. ADMET studies and Molecular docking investigations are performed on the most active hybrid nitroimidazole derivatives 9g and 9k with 4-hydroxytamoxifen (4-OHT) at the human estrogen receptor alpha (hER) during binding active sites to study the ligand–protein interactions and free binding energies at atomic levels. The triazole ring in the 9g derivative forms a hydrogen bond with Asp58 with distance 3.2 Å. And it is found that polar contact with His231 amino acid residue.In silico assessment of the compounds showed very good pharmacokinetic properties based on their physicochemical values, also the ADMET criteria of the most active hybrid systems are within the acceptable range.

Water-Aluminum Interaction as Driving Force of Linde Type A Aluminophosphate Hydration.

Linde type A (LTA) aluminophosphate is a promising candidate for an energy storage material used for low-temperature solar and waste-heat management. The mechanism of reversible water adsorption, which is the basis for potential industrial applications, is still not clear. In this paper, we provide mechanistic insight into various aspects of the hydration process using molecular modeling methods. Building on accurate DFT calculations and available experimental data, we first refine the existing empirical force-field used in subsequent classical molecular dynamics simulations that captures the relevant physics of the water binding process. We succeed in fully reproducing the experimentally determined X-ray structure factors and use them to estimate the number of water molecules present in the fully hydrated state of the material. Furthermore, we show that the translational and orientational mobility of the confined water is significantly reduced and resembles the dynamics of glassy systems.

Rare coordination behavior of triethanolamine ligand in [Cu(H3tea)2](3,4-dimethoxycinnamate)2·2H2O: Synthesis, characterization, single crystal X-ray structure determination, Hirshfeld calculation and molecular docking studies

Herein, unusual and rare coordination behavior of triethanolamine (H3tea) ligand in copper(II) carboxylate complex; [Cu(H3tea)2](3,4-DMC)2·2H2O; DMC = 3,4-dimethoxycinnamate, has been explored and structurally reported for the first time. Synthesis and crystallization of the complex has been achieved at ambient conditions using methanol-water (4:1 v/v) as a solvent mixture after very slow evaporation. Furthermore, the complex is characterized by elemental analyses, spectroscopic methods (UV–Visible, FT-IR) and structure elucidation has been carried out by single crystal X-ray structure analyses. X-ray structure determination of the complex revealed the presence of [Cu(H3tea)2]2+ as a cation and two moieties of 3,4-dimethoxycinnamate as anions forming a second sphere coordination complex. Various weak (non-covalent) interactions such as CH∙∙∙O, and OH∙∙∙O impart an important role in the lattice stabilization of a second sphere coordination complex. Hirshfeld surface calculations clearly revealed the existence of interplay of non-covalent interactions in lattice stabilization. The theoretical calculation also suggested the total interaction energies in this complex, i.e. 32.6 kj.mol−1. In addition, molecular docking investigations of this complex clearly showed that there is substantial protein-ligand interaction with best binding scores of -7.2, -7.3, -7.9 kcal/mol against 4GKU, 3W4P, 5CGX and -5.8, -7.8 kcal/mol against 4HFZ, 1D5R proteins lesser than parent acid i.e. 3,4-dimethoxycinnamic acid indicating the potential of this complex; [Cu(H3tea)2](3,4-DMC)2·2H2O as antibacterial and anticancer agent respectively.

Synthetic Modulation of an Unstable Dehydrosecodine-type Intermediate and Its Encapsulation into a Confined Cavity Enable Its X-ray Crystallographic Observation.

Numerous indole alkaloids such as the iboga- and aspidosperma-type are believed to be biosynthesized via a common hypothetical intermediate, dehydrosecodine. The highly reactive nature of dehydrosecodine-type compounds has hampered their isolation and structural elucidation. In this study, we achieved the first X-ray structural determination of a dehydrosecodine-type compound by integrating synthetic optimization of the reactivity and stabilizing the fragile molecule by encapsulation into a supramolecular host. Formation of a 1 : 1 complex of the dehydrosecodine-type labile guest bearing both vinyl indole and dihydropyridine units with the host was observed. This integrated approach not only provides insights into the biosynthetic conversions but also allows stabilization and storage of the reactive and otherwise short-lived intermediate within the confined hydrophobic cavity.

Crystal structural investigations of heme protein derivatives resulting from reactions of aryl- and alkylhydroxylamines with human hemoglobin

Phenylhydroxylamine (PhNHOH) and nitrosobenzene (PhNO) interact with human tetrameric hemoglobin (Hb) to form the nitrosobenzene adduct Hb(PhNO). These interactions also frequently lead to methemoglobin formation in red blood cells. We utilize UV–vis spectroscopy and X-ray crystallography to identify the primary and secondary products that form when PhNHOH and related alkylhydroxylamines (RNHOH; R = Me, t-Bu) react with human ferric Hb. We show that with MeNHOH, the primary product is Hb[α-FeIII(H2O)][β-FeII(MeNO)], in which nitrosomethane is bound to the β subunit but not the α subunit. Attempts to isolate a nitrosochloramphenicol (CAMNO) adduct resulted in our isolation of a Hb[α-FeII][β-FeII-cySOx]{CAMNO} product (cySOx = oxidized cysteine) in which CAMNO was located outside of the protein in the solvent region between the β2 and α2 subunits of the same tetramer. We also observed that the βcys93 residue had been oxidized. In the case of t-BuNHOH, we demonstrate that the isolated product is the β-hemichrome Hb[α-FeIII(H2O)][β-FeIII(His)2]{t-BuNHOH}, in which the β heme has slipped ∼4.4 Å towards the solvent exterior to accommodate the bis-His heme coordination. When PhNHOH is used, a similar β-hemichrome Hb[α-FeIII(H2O)][β-FeIII(His)2-cySOx]{PhNHOH} was obtained. Our results reveal, for the first time, the X-ray structural determination of a β-hemichrome in a human Hb derivative. Our UV–vis and X-ray crystal structural result reveal that although Hb(PhNO) and Hb(RNO) complexes may form as primary products, attempted isolation of these products by crystallization may result in the structural determination of their secondary products which may contain β-hemichromes en route to further protein degradation.

Completely o-Phenylene Bridged N4-Cyclophane: A Missing Link in the Phenylene Bridge N4-Cyclophane Family

Unprecedented types of metal-free complete o-phenylene bridged N4-cyclophanes (M1 and M2) have been synthesized via sequential palladium-catalyzed Buchwald-Hartwig N-arylation reactions. These cyclophanes may be considered as aromatic analogues of aliphatic group-spaced N4-macrocycles. These have been characterized fully using physicochemical characterization techniques and finally by single crystal X-ray structure determination. Their redox and spectral properties have been characterized by cyclic voltammetry, UV-vis spectro-electrochemistry, fluorescence spectral studies, and DFT calculations. These studies have shown rich redox, spectral, and photophysical properties that could make both M1 and M2 potential candidates for various applications.

Synthesis, single crystal X-ray structure determination, Hirshfeld surface analysis, crystal voids studies, and density functional theory calculations of N-carbamothioylbenzamide and 1,3,5-triazinane-2,4,6-trithione co-crystal

Synthesis, single crystal X-ray structure determination, Hirshfeld surface analysis, crystal voids studies, and density functional theory calculations of N-carbamothioylbenzamide and 1,3,5-triazinane-2,4,6-trithione co-crystal

Synthesis, single crystal X-ray structure determination, Hirshfeld surface analysis, interaction energies, and density functional theory calculations of N,N'-(cyclohexane-1,4-dicarbonothioyl)bis(4-methylbenzamide)

The present research offers convenient synthesis, molecular and crystal structures of C2 symmetric N,N'-(cyclohexane-1,4-dicarbonothioyl)bis(4-methylbenzamide). The synthesized bis-amide crystallized in monoclinic crystal system, having space group of P 21/c with a = 17.8669 (14) Å, b = 6.3495 (5) Å, c = 9.6195 (7) Å, β = 101.567 (8)°, Z = 2 and V = 1069.13 (15) Å3. The asymmetric unit contains cyclohexane ring located on an inversion center, one dicarbonothioyl and one methylbenzamide group. In the crystal structure, the N–H···O, C–H···O and C–H···S hydrogen bonds link the molecules, enclosing R21(6), R22(16) and R22(22) ring motifs, into three-dimensional architecture. Hirshfeld surface (HS) analysis revealed the prevailing significance of hydrogen bond contacts. HS analysis suggested prominent contributions for crystal packing encompasses H…H (49.6%), H…C/C…H (18.9%) and H…S/S…H (16.0%). Hydrogen bonding and van der Waals contacts are the dominant interactions in the crystal packing. The respective volume of the crystal voids and the percentage of free spaces in the unit cell is calculated as 117.87 Å3 and 11.02% indicating the absence of any large cavity within the crystal and the associated stability of the crystal. Evaluation of the electrostatic, dispersion and total energy frameworks revealed that stabilization is dominated by dispersion energy contribution. Moreover, optimized molecular structure, using density functional theory at B3LYP/6-311G(d,p) level, were compared with the experimentally determined ones. HOMO–LUMO energy gap was determined, frequencies and molecular electrostatic potential surface were calculated at the B3LYP/6–311G level to predict sites for electrophilic and nucleophilic attacks. The significant charge polarity, half molecule spread of HOMO and LUMO impart bis-amide nucleophilic as well as electrophilic behavior with relative potency depending upon the nature of the attacking molecule.

(4Z,4′Z)-2,2′-(Ethane-1,2-diylbis(sulfanediyl))bis(1-phenyl)-4-(pyridin-2-ylmethylene)-1H-imidazol-5(4H)-one)dicopper(II) Tetrabromide

The title compound was characterized for the first time by a full range of physical methods including UV, FTIR, mass spectrometry and X-ray structure determination. The copper atoms have a distorted tetrahedral structure close to a trigonal pyramid.

Structures, Characterization and DFT Studies of Four Novel Nickel Phenanthroline Complexes

Four Ni–phen complexes were prepared and characterized by FTIR and thermal analysis. The X-ray structure determination shows that the geometry around NiII ions in complexes 1, [Ni(phen)2(H2O)(ONO2)](NO3); 2, [Ni(phen)2(H2O)Cl]Cl; 3, [Ni(phen)(CH3CN)(ONO2)(O2NO)] and 4, [Ni(phen)3](NO3)2.H2O.C2H5OH is considerably distorted octahedral. The coordination core geometries in 1, 2, 3 and 4 are NiN4O2, NiN4OCl, NiN3O3 and NiN6, respectively. The crystal packing and crystal supramolecularity analysis of the complexes reveal the importance of aryl∙∙∙aryl interactions, and both offset face-to-face (OFF) and edge-to-face (EF) motifs are dominant. Hydrogen bonding interactions in these structures reinforce aryl∙∙∙aryl interactions. FTIR proved the coordination of M-N and M-Cl as well as the presence of nitro groups in the complexes, while thermal analysis revealed that the loss of water, chloro and nitro groups occurred before the degradation of phenanthroline ligand in the complexes. DFT study indicates that a strong correlation exists among theoretical and experimentally determined geometric parameters with distorted octahedral geometry. The charge density in HOMO is localized on the metal halogen bond and nitrate group (NO3−), whereas the LUMO cloud density is mainly distributed on the phen ring, which indicates the electroactive nature of the ring.

Synthesis, molecular structure, mesomorphism and DFT/TD-DFT calculations of square-planar bis[1-((p-tolylimino)methyl)naphthalen-2-olato-κ2N,O]nickel and copper(II) complexes

Reaction of the Schiff base ligand (E)-1-((p-tolylimino)methyl)naphthalen-2-ol (HL) with nickel(II) and copper(II) acetate provides the bis[1-((p-tolylimino)methyl)naphthalen-2-olato-κ2N,O]Ni and Cu(II) complexes (1 and 2), respectively. X-ray structure determinations reveal coordination of two N,O-chelate Schiff base ligands to the metal ion in a square-planar geometry for both the complexes. The phase purity of bulk microcrystalline samples is confirmed by Powder XRD patterns. The solid-state magnetic moment (µeff = 2.73 (1) and 2.06 µB (2)) indicates paramagnetic nature of the complexes. Thermal analyses by differential scanning calorimetry (DSC) and polarizing light microscopic (PLM) observations suggest mesomorphic property for 2. Cyclic voltammograms recommend a quasi-reversible two one electron charge transfer processes for 2 in dimethylformamide. Geometry optimization and excited state properties by DFT/TD-DFT calculations reproduce the experimental bond distances and angles as well as the experimental electronic spectra.

Broad-spectrum coronavirus 3C-like protease peptidomimetic inhibitors effectively block SARS-CoV-2 replication in cells: Design, synthesis, biological evaluation, and X-ray structure determination

Despite the approval of vaccines, monoclonal antibodies and restrictions during the pandemic, the demand for new efficacious and safe antivirals is compelling to boost the therapeutic arsenal against the COVID-19. The viral 3-chymotrypsin-like protease (3CLpro) is an essential enzyme for replication with high homology in the active site across CoVs and variants showing an almost unique specificity for Leu-Gln as P2–P1 residues, allowing the development of broad-spectrum inhibitors.The design, synthesis, biological activity, and cocrystal structural information of newly conceived peptidomimetic covalent reversible inhibitors are herein described. The inhibitors display an aldehyde warhead, a Gln mimetic at P1 and modified P2–P3 residues. Particularly, functionalized proline residues were inserted at P2 to stabilize the β-turn like bioactive conformation, modulating the affinity. The most potent compounds displayed low/sub-nM potency against the 3CLpro of SARS-CoV-2 and MERS-CoV and inhibited viral replication of three human CoVs, i.e. SARS-CoV-2, MERS-CoV, and HCoV 229 in different cell lines. Particularly, derivative 12 exhibited nM-low μM antiviral activity depending on the virus, and the highest selectivity index. Some compounds were co-crystallized with SARS-CoV-2 3CLpro validating our design. Altogether, these results foster future work toward broad-spectrum 3CLpro inhibitors to challenge CoVs related pandemics.