Water-mediated Interactions Research Articles

A combination of cross-neutralizing antibodies synergizes to prevent SARS-CoV-2 and SARS-CoV pseudovirus infection

SummaryCoronaviruses have caused several human epidemics and pandemics including the ongoing coronavirus disease 2019 (COVID-19). Prophylactic vaccines and therapeutic antibodies have already shown striking effectiveness against COVID-19. Nevertheless, concerns remain about antigenic drift in SARS-CoV-2 as well as threats from other sarbecoviruses. Cross-neutralizing antibodies to SARS-related viruses provide opportunities to address such concerns. Here, we report on crystal structures of a cross-neutralizing antibody, CV38-142, in complex with the receptor-binding domains from SARS-CoV-2 and SARS-CoV. Recognition of the N343 glycosylation site and water-mediated interactions facilitate cross-reactivity of CV38-142 to SARS-related viruses, allowing the antibody to accommodate antigenic variation in these viruses. CV38-142 synergizes with other cross-neutralizing antibodies, notably COVA1-16, to enhance neutralization of SARS-CoV and SARS-CoV-2, including circulating variants of concern B.1.1.7 and B.1.351. Overall, this study provides valuable information for vaccine and therapeutic design to address current and future antigenic drift in SARS-CoV-2 and to protect against zoonotic SARS-related coronaviruses.

Cavitand Complexes in Aqueous Solution: Collaborative Experimental and Computational Studies of the Wetting, Assembly, and Function of Nanoscopic Bowls in Water.

Water is the dominant liquid on Earth. Despite this, the main focus of supramolecular chemistry research has been on binding and assembly events in organic solvents. This arose because it is more straightforward to synthesize organic-media-soluble hosts and because of the relative simplicity of organic solvents compared to water. Nature, however, relies on water as a solvent, and spurred by this fact, supramolecular chemists have recently been making forays into the aqueous domain to understand water-mediated non-covalent interactions. These studies can benefit from the substantial understanding of the hydrophobic effect and electrostatic interactions developed by physical chemists. Nearly 20 years ago, the Gibb group first synthesized a class of water-soluble host molecules, the deep-cavity cavitands, that possess non-polar pockets that readily bind non-polar moieties in aqueous solution and are capable of assembling into a wide range of complexes with distinct stoichiometries. As such, these amphipathic host species are ideal platforms for studying the role of negatively curved features on guest complexation and the structural requirements for guided assembly processes driven by the hydrophobic effect. Here we review the collaborative experimental and computational investigations between Gibb and Ashbaugh over the past 10 years exploring questions including the following: How does water wet/solvate the non-polar surfaces of non-polar pockets? How does this wetting control the binding of non-polar guests? How does wetting affect the binding of anionic species? How does the nature and size of a guest size impact the assembly of cavitand hosts into multimeric capsular complexes? What are the conformational motifs of guests packed within the confines of capsular complexes? How might the electrostatic environment engendered by hosts impact the properties and reactivity of internalized guests?

Identification of Potent Reverse Indazole Inhibitors for HPK1.

Hematopoietic progenitor kinase (HPK1), a negative regulator of TCR-mediated T-cell activation, has been recognized as a novel antitumor immunotherapy target. Structural optimization of kinase inhibitor 4 through a systematic two-dimensional diversity screen of pyrazolopyridines led to the identification of potent and selective compounds. Crystallographic studies with HPK1 revealed a favorable water-mediated interaction with Asp155 and a salt bridge to Asp101 with optimized heterocyclic solvent fronts that were critical for enhanced potency and selectivity. Computational studies of model systems revealed differences in torsional profiles that allowed for these beneficial protein-ligand interactions. Further optimization of molecular properties led to identification of potent and selective reverse indazole inhibitor 36 that inhibited phosphorylation of adaptor protein SLP76 in human PBMC and exhibited low clearance with notable bioavailability in in vivo rat studies.

Second distinct conformation of the phosphohistidine loop in succinyl-CoA synthetase.

Succinyl-CoA synthetase (SCS) catalyzes a reversible reaction that is the only substrate-level phosphorylation in the citric acid cycle. One of the essential steps for the transfer of the phosphoryl group involves the movement of the phosphohistidine loop between active site I, where CoA, succinate and phosphate bind, and active site II, where the nucleotide binds. Here, the first crystal structure of SCS revealing the conformation of the phosphohistidine loop in site II of the porcine GTP-specific enzyme is presented. The phosphoryl transfer bridges a distance of 29 Å between the binding sites for phosphohistidine in site I and site II, so these crystal structures support the proposed mechanism of catalysis by SCS. In addition, a second succinate-binding site was discovered at the interface between the α- and β-subunits of SCS, and another magnesium ion was found that interacts with the side chains of Glu141β and Glu204β via water-mediated interactions. These glutamate residues interact with the active-site histidine residue when it is bound in site II.

Hygroscopicity of Hofmeister Salts and Glycine Aerosols-Salt Specific Interactions.

The Hofmeister effect of inorganic ions to precipitate proteins has been used to understand the coagulation phenomenon in colloid and protein science. Herein, for the first time, this effect is studied on the hygroscopicity of aerosols using ATR-FTIR spectroscopy. The representative Hofmeister salts (MgSO4, KCl, NH4NO3) and amino acid (glycine) with different amino acid/salt molar ratios (ASRs) are mixed and atomized into micrometer-sized particles. For mixed kosmotrope (MgSO4)/glycine and chaotrope (NH4NO3)/glycine with an ASR of 1:1, both ERHs (efflorescence relative humidities) and DRHs (deliquescence relative humidities) are absent. However, for the mixtures of glycine and neutral salt (KCl), no DRH is observed while 66.2 and 61.4% ERH of glycine is detected for mixtures with ASRs of 1:1 and 1:3, respectively, which is similar to pure glycine. For the mixture of NH4NO3/glycine with an ASR of 1:3, ERH and DRH are found to be 15.4 and 32.2% RH, less than that of pure NH4NO3. Further, interactions between glycine-salt and/or water is also studied in the mixtures during hydration and dehydration. Water-mediated ion-glycine interaction is detected based on the two glycine bands merging into one band. Glycine-SO42- interaction is present for glycine/sulfate in all ASRs, while glycine-NO3- interaction is only seen for 1:3 glycine/NH4NO3 mixtures during hydration. This work opens a window to understand the Hofmeister effect on the hygroscopicity of atmospheric aerosols.

Capturing Water Networks during Ligand Binding with the Site-Identification by Ligand Competitive Saturation-Monte Carlo Approach

Water molecules impact interactions between proteins and their ligands, making a significant contribution to ligand binding orientation and affinity. Water molecules can significantly affect the energetics of ligand binding in a favorable or unfavorable manner associated, in part, with the energetic penalty for displacing waters that occupy a binding site. In computer-aided drug design, the challenge is to calculate water mediated interactions and their energetics between the ligand and the protein. In this work, an extension of the Site Identification by Ligand Competitive Saturation-Monte Carlo (SILCS-MC) docking approach is presented to determine the positioning of the water molecules in the binding site and estimate the energetic contribution of water to ligand binding. We employ this methodology on variety of protein targets where water mediated interactions between the protein and ligands plays an important role. The efficacy of this approach is reflected in rank-ordering ligand affinities, binding affinities predictions, and structure orientation of the ligands. The SILCS methodology is of utility for characterization of functional group affinities for the entire 3D region encompassed by a protein or other macromolecule. This approach is based on sampling the conformational space of multiple solutes representing various functional groups in the presence of explicit water of a protein target using a combination of oscillating chemical potential Grand Canonical Monte Carlo and Molecular Dynamics simulations. The presented approach offers new possibilities in revealing water networks and their contributions to the binding affinity of a ligand to a protein.

Spectroscopic and Structural Characterization of Water-Shared Ion-Pairs in Aqueous Sodium and Lithium Hydroxide.

The structures of the ion-pairs formed in aqueous NaOH and LiOH solutions are elucidated by combining Raman multivariate curve resolution (Raman-MCR) experiments and ab initio molecular dynamics (AIMD) simulations. The results extend prior findings to reveal that the initially formed ion-pairs are predominantly water-shared, with the hydroxide ion retaining its full first hydration-shell, while direct contact ion-pairing only becomes significant at higher concentrations. Our results confirm previous experiments and simulations indicating greater ion-pairing in aqueous LiOH than NaOH as well as at high temperatures. Our results further imply that NaOH and LiOH ion-pairing free energies have an approximately linear (rather than square-root) dependence on ion concentration (in the molar range), with positive enthalpies and entropies that increase with concentration, thus implying that water-mediated interactions enthalpically disfavor and entropically favor ion-pair formation.

Water-Mediated Interactions Determine Helix Formation of Peptides in Open Nanotubes.

Water-mediated interactions (WMIs) play diverse roles in molecular biology. They are particularly relevant in geometrically confined spaces such as the interior of the chaperonin, at the interface between ligands and their binding partners, and in the ribosome tunnel. Inspired in part by the geometry of the ribosome tunnel, we consider confinement effects on the stability of peptides. We describe results from replica exchange molecular dynamics simulations of a system containing a 23-alanine or 23-serine polypeptide confined to nonpolar and polar nanotubes in the gas phase and when open to a water reservoir. We quantify the effect of water in determining the preferred conformational states of these polypeptides by calculating the difference in the solvation free energy for the helix and coil states in the open nanotube in the two phases. Our simulations reveal several possibilities. We find that nanoscopic confinement preferentially stabilizes the helical state of polypeptides with hydrophobic side chains, which is explained by the entropic stabilization mechanism proposed on the basis of polymer physics. Polypeptide chains with hydrophilic side chains can adopt helical structures within nanotubes, but helix formation is sensitive to the nature of the nanotube due to WMIs. We elaborate on the potential implications of our findings to the stability of peptides in the ribosome tunnel.

Water-Mediated Attractive Interaction between Negatively Charged GO Nanosheets at the Air–Water Interface

Understanding the interaction of ionized two-dimensional (2D) graphene oxide (GO) nanosheets at the air–water interface can improve the self-assembly framework of GO sheets. Herein, the potential o...

Behavior of water confined between hydrophobic surfaces with grafted segments

Many naturally occurring as well as functionalized interfaces are covered with protruding segments or side-chain moieties. These protruding segments influence interfacial properties, and play an important role in diverse physical processes that are understood to be driven by water-mediated interactions, ranging from protein folding, protein dimerization to nanoparticle self-assembly in aqueous media. We have studied the behavior of water confined between two planar hydrophobic surfaces with grafted segments using molecular simulations. We show the presence of these segments makes the confined region significantly more hydrophobic. By systematically varying the flexibility of these segments over a range spanning two orders of magnitude for two different grafting densities, we report that the flexibility of the segments itself has no effect on the free energy profile of confined water. We also find that the evaporation free energy barrier increases and the condensation free energy barrier decreases as the density of the grafted segment is increased.

The molecular mechanism behind the stabilization of insulin by choline and geranate (CAGE) ionic liquids - computational insights into oral insulin drug formulation.

Insulin is a principal hormone that is involved in the regulation of glucose levels in the blood. Oral insulin formulation is a recent development in drug delivery systems. Biocompatible choline-based ionic liquids (ILs) show promising antibacterial activity and are useful for oral and transdermal drug delivery applications. Choline and geranate (CAGE) ILs enhance the stability and oral efficacy of insulin delivery. The molecular mechanism behind insulin formulation in the oral form is at issue. In the present work, the molecular-level understanding of CAGE ILs in insulin is scrutinized by employing atomistic molecular dynamics (MD) simulations. To identify the stability of insulin in an IL medium, we have studied a series of concentration (mole fraction 0.05-1.00) of CAGE ILs with an insulin dimer. It can be well evidenced from the experimental reports that in an aqueous medium, there is a refashioning of CAGE nanostructures at 0.50 mole fraction. It is found from our calculations that the first solvation shell of insulin is readily occupied by choline and geranate ions in the presence of water. Moreover, the geranate ions strongly interacted with the water molecules and thereby, eliminating the intermolecular hydrogen bonding (H-bonding) interactions towards the insulin at 0.30-0.50 mole fraction of CAGE ILs. The most desirable 0.30-0.50 mole fraction of CAGE invigorates water-mediated H-bonding interactions with geranate ions, which also enhances the electrostatic behavior around the vicinity of the insulin dimer. These important findings can help in the development of oral insulin drug delivery and related applications.

In silico mutational analyses reveal different ligand-binding abilities of double pockets of medaka fish taste receptor type 1 essential for efficient taste recognition.

Taste receptors are important sensors for the detection of nutrient concentrations in animals. Tastes are recognized by interactions between chemical substances and taste receptors. Recently, the high-resolution X-ray crystal structure of the extracellular ligand-binding domains (LBDs) of medaka fish (Oryzias latipes) taste receptor type 1 (T1r) complexed with ligands (amino acids) was determined. Medaka fish T1r is a heterodimer composed of two different LBDs, T1r2aLBD and T1r3LBD. In this study, we performed all-atom molecular dynamics (MD) simulations on this heterodimer (T1r2aLBD-T1r3LBD) to address mutational effects on key residues near each ligand-binding pocket in recognizing one of the ligands (L-Gln). For T1r2aLBD, Ser165 is important in ligand recognition due to its direct hydrogen bonding with the ligand. After mutating Ser165 to Ile or Ala, the direct hydrogen bonds between the ligand and the binding pocket were weakened, which destabilized the ligand-binding form of T1r2aLBD. For T1r3LBD, Ser300 is important in ligand recognition. The water-mediated hydrogen bond with the side-chain hydroxyl group of Ser300 is a single interaction that maintains the ligand-binding form of T1r3LBD. After mutating Ser300 to Glu or Ala, both mutant systems almost maintained their ligand-binding form. As a mechanism for maintaining the binding form of T1r3LBD, alternative hydrogen bonds were formed as direct interactions instead of the indirect water-mediated interactions found in the wild-type system, which stabilized the binding form of T1r3LBD. Judging from our in silico mutational analyses, T1r2aLBD was structurally destabilized by the amino acid mutations. Therefore, it might be required that the ligand-binding pocket of T1r2aLBD is composed of a set of specific residues to maintain its ligand-binding form. On the contrary, T1r3LBD was robust enough to withstand the amino acid mutations. These different ligand-binding abilities of both LBDs provide multiple binding modes, which might be helpful for discriminating various taste substances or detecting concentrations of nutrients efficiently.

Nanoscale Lubrication in Biosystems as Rationalized in Terms of Fractons and Spectral-Mechanical Properties of Networked Biopolymers in Ionic Solutions

Articular cartilage is a natural tribochemical device just-designed by nature. Yet, a vivid debate goes on toward the mechanisms by which its nanoscopic properties facilitate lubrication in terms of ultralow static and kinetic friction coefficients. In this concisely conducted conceptual discussion, we wish to point out that a nanoscale tribomechanistic description based upon certain viscoelastic quanta, called fractons, expressing spectral-mechanical properties of nets under the influence of force/pressure factor(s), may contribute substantially to the elucidation of ultralow coefficients of friction in the articular cartilage of predictable relaxational response. Our example unveils a part of a mechanically responsive network, such as a tied piece of hyaluronan molecule, fit in an Edwards type tube, in which upon water-mediated interaction of lipids with the hyaluronan when subjected to loading at the nanoscale, a consecutive stress-field and ion diffusion action is going to occur simultaneously. It results in a natural-logarithmic formula that interrelates a number of hyaluronans interactive residues, N, with certain molecular-elastic (an exponent{gamma} ) and surface-to-volume (nano-colloid type) characteristics of around 2/3 to emerge near thermodynamic equilibrium, that is to say after a frictional loading action performed. It enables to relate uniquely a value of the exponent 0 ] 1/3 from above for large enough N -s. A periodic vibrational super-biopolymers mode is exploited, leading to a change in the nanoscale friction-lubrication period from which an opportunity to involve an essential contribution to the (nanoscale) coefficient of friction arises. PACS numbers71.10.+x, 81.30.Fb, 05.70.Fh, 05.60.+w

Nonreciprocal interactions induced by water in confinement

Water mediates electrostatic interactions via the orientation of its dipoles around ions, molecules, and interfaces. This induced water polarization consequently influences multiple phenomena. In particular, water polarization modulated by nanoconfinement affects ion adsorption and transport, biomolecular self-assembly, and surface chemical reactions. Therefore, it is of paramount importance to understand how water-mediated interactions change at the nanoscale. Here we show that near the graphene surface anion-cation interactions do not obey the translational and isotropic symmetries of Coulomb's law. We identify a new property, referred to as non-reciprocity, which describes the non-equivalent and directional interaction between two oppositely charged ions near the confining surface when their positions with respect to the interface are exchanged. Specifically, upon exchange of the two ions' positions along the surface normal direction the interaction energy changes by about 5$k_BT$. In both cases, confinement enhances the attraction between two oppositely charged ions near the graphene surface, while intercalation of one ion into the graphene layers shifts the interaction to repulsive. While the water permittivity in confinement is different from that in bulk, the effects observed here via molecular dynamics simulations and X-ray reflectivity experiments cannot be accounted for by current permittivity models. Our work shows that the water structure is not enough to infer electrostatic interactions near interfaces.

Crystallographic Snapshots of the Dunathan and Quinonoid Intermediates provide Insights into the Reaction Mechanism of Group II Decarboxylases

PLP-dependent enzymes catalyze a plethora of chemical reactions affecting diverse physiological functions. Here we report the structural determinants of the reaction mechanism in a Group II PLP-dependent decarboxylase by assigning two early intermediates. The in-crystallo complexes of the PLP bound form, and the Dunathan and quinonoid intermediates, allowed direct observation of the active site interactions. The structures reveal that a subtle rearrangement of a conserved Arg residue in concert with a water-mediated interaction with the carboxylate of the Dunathan intermediate, appears to directly stabilize the alignment and facilitate the release of CO2 to yield the quinonoid. Modeling indicates that the conformational change of a dynamic catalytic loop to a closed form controls a conserved network of hydrogen bond interactions between catalytic residues to protonate the quinonoid. Our results provide a structural framework to elucidate mechanistic roles of residues that govern reaction specificity and catalysis in PLP-dependent decarboxylation.

Long-Range Ordered Water Correlations between A–T/C–G Nucleotides

Summary The interactions between complementary base pairs are crucial to the helical duplex structure of DNA. Although base-base interactions have been widely reported, the study of long-range interactions in the base-pairing processes is still a major challenge in this field. Here, we first study the long-range interactions between the base pairs by atomic force microscopy-based single-molecule force spectroscopy (SMFS). The SMFS results of A–T/C–G imply that there are weak long-range interactions between them, which have a range of 15–25 nm. Raman spectroscopy results imply that these weak interactions can be attributed to multiplex hydrogen bond interaction of ordered water structure between A–T/C–G nucleotides. In addition, the theoretical calculations deduce that the ideal structure of these water molecules exhibits a specific helical structure. Our findings might open up a new understanding of biological assembly processes and provide helpful strategies for bio-nanotechnology.

Structure of cyclin-dependent kinase 2 (CDK2) in complex with the specific and potent inhibitor CVT-313.

CVT-313 is a potent CDK2 inhibitor that was identified by screening a purine-analogue library and is currently in preclinical studies. Since this molecule has the potential to be developed as a CDK2 inhibitor for cancer therapy, the potency of CVT-313 to bind and stabilize CDK2 was evaluated, together with its ability to inhibit aberrant cell proliferation. CVT-313 increased the melting temperature of CDK2 by 7°C in thermal stabilization studies, thus indicating its protein-stabilizing effect. CVT-313 inhibited the growth of human lung carcinoma cell line A549 in a dose-dependent manner, with an IC50 of 1.2 µM, which is in line with the reported biochemical potency of 0.5 µM. To support the further chemical modification of CVT-313 and to improve its biochemical and cellular potency, a crystal structure was elucidated in order to understand the molecular interaction of CVT-313 and CDK2. The crystal structure of CDK2 bound to CVT-313 was determined to a resolution of 1.74 Å and clearly demonstrated that CVT-313 binds in the ATP-binding pocket, interacting with Leu83, Asp86 and Asp145 directly, and the binding was further stabilized by a water-mediated interaction with Asn132. Based on the crystal structure, further modifications of CVT-313 are proposed to provide additional interactions with CDK2 in the active site, which may significantly increase the biochemical and cellular potency of CVT-313.

Characterization of active/binding site residues of peptidyl-tRNA hydrolase using biophysical and computational studies

All mRNAs cannot be translated into full-length proteins due to ribosome-stalling that leads to release of peptidyl-tRNA which can be lethal for bacterial survival. The enzyme peptidyl-tRNA hydrolase (PtH) hydrolyses the ester bond between nascent peptide and tRNA of peptidyl-tRNA and rescues the cells from toxicity. PtH is an essential enzyme in bacteria and inhibiting this crucial enzyme can serve to combat bacterial diseases. But due to lack of understanding about the catalytic mechanism of PtH, its inhibitors have not been developed. In this work, we have carried out the binding studies of M. tuberculosis and E. coli PtH with the peptidyl-tRNA analogue (puromycin) using ITC, FTIR, CD experiments followed by docking and MD simulations to identify the potential active site residues that would help to design PtH inhibitors. Binding studies of puromycin with both PtH by ITC experiments demonstrate similar thermodynamic parameters and three fold difference in their KD. CD and FTIR studies detected changes in secondary structure composition of PtH in the presence of puromycin with different degree of perturbation. Though interactions with puromycin are conserved in both proteins, modelling studies revealed that water mediated interactions in M. tb-PtH resulting in higher affinity to puromycin.

An Evolutionary Marker of the Ribokinase Superfamily Is Responsible for Zinc-Mediated Regulation of Human Pyridoxal Kinase

The ribokinase superfamily catalyzes the phosphorylation of a vast diversity of substrates, and its members are characterized by the conservation of a common structural fold along with highly conserved sequence motifs responsible for phosphoryl transfer (GXGD) and stabilization of the metal-nucleotide complex (NXXE). Recently, a third motif (HXE) exclusive from ADP-dependent enzymes was identified, with its glutamic acid participating in water-mediated interactions with the metal-nucleotide complex and in stabilization of the ternary complex during catalysis. In this work, we bioinformatically determine that the aspartic acid of another motif (DPV), exclusively found in hydroxyethyl thiazole (THZK), hydroxymethyl pyrimidine (HMPK) and pyridoxal kinases (PLK), is structurally equivalent to the acidic residue in the HXE motif. Moreover, this residue is highly conserved among all ribokinase superfamily members. To determine whether the functional role of the DPV motif is similar to the HXE motif, we employed molecular dynamics simulations using crystal structures of phosphoryl donor substrate-complexed THZK and PLK, showing that its aspartic acid participated in water-mediated or direct interactions with the divalent metal of the metal-nucleotide complex. Lastly, enzyme kinetic assays on human PLK, an enzyme that utilizes zinc, showed that site-directed mutagenesis of the aspartic acid from the DPV motif abolishes the inhibition of this enzyme by increasing free zinc concentrations. Altogether, our results highlight that the DPV and HXE motifs are evolutionary markers of the functional and structural divergence of the ribokinase superfamily and evidence the role of the DPV motif in the interaction with both free and nucleotide-complexed divalent metals in the binding site of these enzymes.

PH-Dependent adsorption of aromatic compounds on graphene oxide: An experimental, molecular dynamics simulation and density functional theory investigation

This work provides a comprehensive understanding for the pH-dependent adsorption of aromatic compounds (ACs) on graphene oxide (GO). Isothermal and kinetics experiments indicated both adsorption capacity and adsorption rate were suppressed at higher pH, and the mechanisms were revealed by molecular dynamics simulations and density functional theory calculations. More specifically, π-π, hydrogen bond, vdWs, and water-mediated steric hindrance interactions were examined to reveal how pH affected the adsorption capacity, and microscopic dynamic adsorption process was captured to reveal how pH affected the adsorption rate. Results showed the reduced adsorption capacity at higher pH was mediated by increased electrostatic repulsion, weakened π-π interaction, and increased water-mediated steric hindrance. The pH-dependent behaviour of GO was responsible for the effect of pH on adsorption rate. Self-aggregation of GO at lower pH helped to capture ACs and created more favourable adsorption sites. Upon the adsorption of ACs on GO, GO/water/AC/water/GO sandwich-like structure formed, which was also mediated by solution pH. Overall, pH affects the adsorption of ACs on GO by regulating driving forces, adsorption process, and the configuration property of GOAC complex.