Unravelling the Time Scale of Conformational Plasticity and Allostery in Glycan Recognition by Human Galectin-1

The interaction of human galectin-1 with a variety of oligosaccharides, from di-(N-acetyllactosamine) to tetra-saccharides (blood B type-II antigen) has been scrutinized by using a combined approach of different NMR experiments, molecular dynamics (MD) simulations, and isothermal titration calorimetry. Ligand- and receptor-based NMR experiments assisted by computational methods allowed proposing three-dimensional structures for the different complexes, which explained the lack of enthalpy gain when increasing the chemical complexity of the glycan. Interestingly, and independently of the glycan ligand, the entropy term does not oppose the binding event, a rather unusual feature for protein-sugar interactions. CLEANEX-PM and relaxation dispersion experiments revealed that sugar binding affected residues far from the binding site and described significant changes in the dynamics of the protein. In particular, motions in the microsecond-millisecond timescale in residues at the protein dimer interface were identified in the presence of high affinity ligands. The dynamic process was further explored by extensive MD simulations, which provided additional support for the existence of allostery in glycan recognition by human galectin-1.     

Highly effective acyclic carbohydrate receptors consisting of aminopyridine, imidazole, and indole recognition units

Neutral imidazole/aminopyridine- and indole/aminopyridine-based receptors, 1 and 2, have been established as highly effective and selective carbohydrate receptors. These receptors effectively recognise neutral carbohydrates through multiple interactions, including neutral hydrogen bonds and CH...pi interactions between the sugar CH groups and the aromatic rings of the receptors. The design of these receptors was inspired by the binding motifs observed in the crystal structures of protein-carbohydrate complexes. The formation of very strong complexes with beta-glucopyranoside 5, beta-maltoside 8, and alpha-maltoside 9 in organic media has been characterised by 1H NMR spectroscopy and confirmed by a second, independent technique, namely fluorescence spectroscopy. The syntheses, molecular-modelling studies, binding properties of the receptors 1 and 2 toward selected mono- and disaccharides as well as comparative binding studies with receptors 3 and 4 are described.     

Aromatic-carbohydrate interactions: an NMR and computational study of model systems

The interactions of simple carbohydrates with aromatic moieties have been investigated experimentally by NMR spectroscopy. The analysis of the changes in the chemical shifts of the sugar proton signals induced upon addition of aromatic entities has been interpreted in terms of interaction geometries. Phenol and aromatic amino acids (phenylalanine, tyrosine, tryptophan) have been used. The observed sugar-aromatic interactions depend on the chemical nature of the sugar, and thus on the stereochemistries of the different carbon atoms, and also on the solvent. A preliminary study of the solvation state of a model monosaccharide (methyl beta-galactopyranoside) in aqueous solution, both alone and in the presence of benzene and phenol, has also been carried out by monitoring of intermolecular homonuclear solvent-sugar and aromatic-sugar NOEs. These experimental results have been compared with those obtained by density functional theory methods and molecular mechanics calculations.     

Novel C3-symmetric molecular scaffolds with potential facial differentiation

The conversion of 1,3,5-substituted benzene and mesitylene by electrophilic aromatic substitution and Sonogashira cross-coupling, respectively, furnished the C3-symmetric, hexasubstituted benzene derivatives 1 and 2 with an alternating substitution pattern. Based on the molecular scaffolds obtained, the two systems serve as model compounds for novel receptor molecules with distinct geometric features. X-ray structures have been obtained for 1 and 2, which are discussed in regard to their aptitude as receptor platforms or supramolecular building blocks. By looking at the rotational barriers for the functional groups placed around the molecular scaffolds by variable temperature 1H NMR spectroscopy, 1 and 2 turn out to exist in rapidly interconverting conformations. The alignment of these potential binding groups around the molecular scaffolds should be strongly biased by specific interactions with suitable guest molecules.     

Aromatic Rings as Molecular Determinants for the Molecular Recognition of Protein Kinase Inhibitors

Protein kinases are key enzymes in many signal transduction pathways, and play a crucial role in cellular proliferation, differentiation, and various cell regulatory processes. However, aberrant function of kinases has been associated with cancers and many other diseases. Consequently, competitive inhibition of the ATP binding site of protein kinases has emerged as an effective means of curing these diseases. Over the past three decades, thousands of protein kinase inhibitors (PKIs) with varying molecular frames have been developed. Large-scale data mining of the Protein Data Bank resulted in a database of 2139 non-redundant high-resolution X-ray crystal structures of PKIs bound to protein kinases. This provided us with a unique opportunity to study molecular determinants for the molecular recognition of PKIs. A chemoinformatic analysis of 2139 PKIs resulted in findings that PKIs are “flat” molecules with high aromatic ring counts and low fractions of sp³ carbon. All but one PKI possessed one or more aromatic rings. More importantly, it was found that the average weighted hydrogen bond count is inversely proportional to the number of aromatic rings. Based on this linear relationship, we put forward the exchange rule of hydrogen bonding interactions and non-bonded π-interactions. Specifically, a loss of binding affinity caused by a decrease in hydrogen bonding interactions is compensated by a gain in binding affinity acquired by an increase in aromatic ring-originated non-bonded interactions (i.e., π–π stacking interactions, CH–π interactions, cation–π interactions, etc.), and vice versa. The very existence of this inverse relationship strongly suggests that both hydrogen bonding and aromatic ring-originated non-bonded interactions are responsible for the molecular recognition of PKIs. As an illustration, two representative PKI–kinase complexes were employed to examine the relative importance of different modes of non-bonded interactions for the molecular recognition of PKIs. For this purpose, two FDA-approved PKI drugs, ibrutinib and lenvatinib, were chosen. The binding pockets of both PKIs were thoroughly examined to identify all non-bonded intermolecular interactions. Subsequently, the strengths of interaction energies between ibrutinib and its interacting residues in tyrosine kinase BTK were quantified by means of the double hybrid DFT method B2PLYP. The resulting energetics for the binding of ibrutinib in tyrosine kinase BTK showed that CH–π interactions and π–π stacking interactions between aromatic rings of the drug and hydrophobic residues in its binding pocket dominate the binding interactions. Thus, this work establishes that, in addition to hydrogen bonding, aromatic rings function as important molecular determinants for the molecular recognition of PKIs. In conclusion, our findings support the following pharmacophore model for ATP-competitive kinase inhibitors: a small molecule features a scaffold of one or more aromatic rings which is linked with one or more hydrophilic functional groups. The former has the structural role of acting as a scaffold and the functional role of participating in aromatic ring-originated non-bonded interactions with multiple hydrophobic regions in the ATP binding pocket of kinases. The latter ensure water solubility and form hydrogen bonds with the hinge region and other hydrophilic residues of the ATP binding pocket.     

Systematic Dissection of an Aminopyrrolic Cage Receptor for β‐Glucopyranosides Reveals the Essentials for Effective Recognition

A set of structures designed for the recognition of glucosides has been obtained by systematically destructuring a tripodal aminopyrrolic cage receptor that selectively recognizes octyl‐β‐D ‐glucopyranoside (OctβGlc). NMR spectroscopy and isothermal titration calorimetry binding measurements showed that cleavage of one pillar of the cage was beneficial to the binding properties of the receptor, as long as two residual amino groups of the cleaved pillar were present. Removal of these two residual amino groups produced a dramatic loss of affinity for OctβGlc of the resulting monocyclic analogue of the parent cage receptor. A significant improvement in the binding ability was achieved by replacing one pillar with two aminopyrrolic hydrogen‐bonding arms, despite the loss of a preorganized structure. In contrast to the cage receptor, recognition of OctβGlc was observed, even in a competitive medium (30 % DMF in chloroform). Structural studies in solution, carried out through NMR spectroscopy and molecular modeling calculations, led to the elucidation of the 3D binding modes of the side‐armed monocyclic receptors; this highlighted the key role of the amino groups and demonstrated the occurrence of a rotaxane‐like complex, which featured the octyl chain of the glucoside threaded through the macrocyclic ring.     

Influence of the methyl position on the binding of 5-epi-taiwaniaquinone G to HHb investigated by spectrofluorimetry and molecular modelling

The interactions of human haemoglobin (HHb) with 5-epi-taiwaniaquinone G (G1) and one structural analogue of 5-epi-taiwaniaquinone G (G2) systematically by UV–vis absorption spectroscopy and fluorescence spectroscopy in combination with molecular modelling are studied in this paper. They caused the fluorescence quenching of HHb by the formation of complex. The binding constants and thermodynamic parameters were obtained. The hydrophobic and electrostatic interactions were the predominant intermolecular forces to stabilise these complexes. Results of thermodynamic analysis and molecular modelling showed that G2 was the stronger quencher and bound to HHb with higher affinity. The result of molecular modelling is close to that obtained by experimental methods.     

Structure-activity relationships of GHRP-6 azapeptide ligands of the CD36 scavenger receptor by solid-phase submonomer azapeptide synthesis

The cluster of differentiation 36 (CD36) class B scavenger receptor binds a variety of biologically endogenous ligands in addition to synthetic peptides (i.e., growth hormone-releasing peptides, GHRPs), which modulate biological function related to anti-angiogenic and anti-atherosclerotic activities. Affinity labeling had previously shown that GHRP-6 analogues such as hexarelin, [2-Me-W(2)]GHRP-6 (1), bind to the lysine-rich domain of the CD36 receptor. Moreover, the azapeptide analogue [aza-F(4)]GHRP-6, 2, exhibited a characteristic β-turn conformation as described by CD and NMR spectroscopy and a slightly higher CD36 binding affinity relative to hexarelin (1.34 and 2.37 μM, respectively), suggesting receptor binding was mediated by the conformation and the aromatic residues of these peptide sequences. Ligand-receptor binding interactions were thus explored using azapeptides to examine influences of side-chain diversity and backbone conformation. In particular, considering that aromatic cation interactions may contribute to binding affinity, we have explored the potential of introducing salt bridges to furnish GHRP-6 azapeptide ligands of the CD36 receptor. Fifteen aza-glutamic acid analogues related to 2 were prepared by submonomer solid-phase synthesis. The azapeptide side chains were installed by novel approaches featuring alkylation of resin-bound semicarbazone with Michael acceptors and activated allylic acetates in the presence of phosphazene base (BTPP). Moreover, certain Michael adducts underwent intramolecular cyclization during semicarbazone deprotection, leading to novel pyrrazoline and aza-pyroglutamate N-terminal residues. Structural studies indicated that contingent on sequence the [aza-Glu]GHRP-6 analogues exhibited CD spectra characteristic of random coil, polyproline type II and β-turn secondary structures in aqueous media. In covalent competition binding studies with the GHRP-6 prototype hexarelin bearing a radiotracer, certain [aza-Glu]GHRP-6 azapeptides retained relatively high (2-27 μM) affinity for the CD36 scavenger receptor.     

Impact of planarity of unfused aromatic molecules on G-quadruplex binding: learning from isaindigotone derivatives

G-quadruplex structures are a new class of attractive targets for DNA-interactive anticancer agents. The primary building block of this structure is the G-quartet, which is composed of four coplanar guanines and serves as the major binding site for small molecules. NMR studies and molecular dynamics simulations have suggested that the planarity of G-quartet surface has been highly dynamic in solution. To better investigate how the planarity of unfused aromatic ligand impacts on its quadruplex binding properties, a variety of planarity controllable isaindigotone derivatives were designed and synthesized. The interaction of G-quadruplex DNA with these designed ligands was systematically explored using a series of biophysical studies. The FRET-melting, SPR, and CD spectroscopy results showed that reducing the planarity of their unfused aromatic core resulted in their decreased binding affinity and stabilization ability for G-quadruplex. NMR studies also suggested that these compounds could stack on the G-quartet surface. Such results are in parallel with subsequent molecular modeling studies. A detailed binding energy analysis indicated that van der Waals energy (ΔE(vdw)) and entropy (TΔS) are responsible for their decreased quadruplex binding and stabilization effect. All these results provided insight information about how quadruplex recognition could be controlled by adjusting the planarity of ligands, which shed light on further development of unfused aromatic molecules as optimal G-quadruplex binding ligands.     

Structures of carbohydrate-boronic acid complexes determined by NMR and molecular modelling in aqueous alkaline media

The structures of thermodynamically stable aromatic boronic acid : cyclic carbohydrate chelates in aqueous alkaline media have been studied using 1H NMR spectroscopy and molecular modelling. It is found that interacting saccharides must necessarily possess a synperiplanar diol functionality for complexation to occur. While this is possible for furanose structures which tend to have a puckered planar geometry, for pyranose forms it is postulated that bis-complexation occurs with twist conformers of the pyranose ring, providing the ring has the requisite 1,2 : 3,4 polyol stereochemistry; specifically axial,equatorial : equatorial,axial or equatorial,axial : axial,equatorial orientations. In this respect it is possible to be predictive with regard to individual carbohydrate boronic acid interactions and to give reasonably comprehensive structural assignments to complexed components. In this paper twenty four polyhydroxy compounds have been screened using 1H NMR to monitor complexation along with computational techniques on a model system to substantiate proposed structures. It has been found that all of these materials interact with aromatic mono boronic acids as expected and structures for the resulting chelates are proposed.     

分子力学和分子动力学方法研究不同变质程度烟煤的分子结构

采用分子力学和分子动力学模拟方法 ,研究了不同变质程度烟煤的三维分子构型和能量参数。结果表明 :随变质程度的增加 ,煤分子内平行的芳香片层结构增大。不同变质程度烟煤分子在团聚前后的成键相互作用能仅略有变化 ,其中扭转能Et 的变化相对较为显著 ,并且扭转能随煤阶的增加呈逐渐减小的趋势。非成键作用能 ,特别是超过三个原子的范德华作用能 ,是煤中的重要相互作用能 ,是模型分子团聚的重要驱动力 ,对于烟煤分子聚集状态的形成起着决定性的作用。同时随煤阶的升高 ,超过三个原子的范德华作用能逐渐增加。模型分子的总势能随煤阶的变化呈两头高、中间低的趋势 ,与煤的一些宏观物理性质有一定的相一致性。     

Design, synthesis, and computational studies of inhibitors of Bcl-XL

One of the primary objectives in the design of protein inhibitors is to shape the three-dimensional structures of small molecules to be complementary to the binding site of a target protein. In the course of our efforts to discover potent inhibitors of Bcl-2 family proteins, we found a unique folded conformation adopted by tethered aromatic groups in the ligand that significantly enhanced binding affinity to Bcl-XL. This finding led us to design compounds that were biased by nonbonding interactions present in a urea tether to adopt this bioactive, folded motif. To characterize the key interactions that induce the desired conformational bias, a series of substituted N,N'-diarylureas were prepared and analyzed using X-ray crystallography and quantum mechanical calculations. Stabilizing pi-stacking interactions and destabilizing steric interactions were predicted to work in concert in two of the substitution patterns to promote the bioactive conformation as a global energy minimum and result in a high target binding affinity. Conversely, intramolecular hydrogen bonding present in the third substitution motif promotes a less active, extended conformer as the energetically favored geometry. These findings were corroborated when the inhibition constant of binding to Bcl-XL was determined for fully elaborated analogues bearing these structural motifs. Finally, we obtained the NMR solution structure of the disubstituted N,N'-diarylurea bound to Bcl-XL demonstrating the folded conformation of the urea motif engaged in extensive pi-interactions with the protein.     

A Water‐Soluble Perylene Bisimide Cyclophane as a Molecular Probe for the Recognition of Aromatic Alkaloids

Herein, we report a water‐soluble macrocyclic host based on perylene bisimide (PBI) chromophores that recognizes natural aromatic alkaloids in aqueous media by intercalating them into its hydrophobic cavity. The host–guest binding properties of our newly designed receptor with several alkaloids were studied by UV/Vis and fluorescence titration experiments as the optical properties of the chromophoric host change significantly upon complexation of guests. Structural information on the host–guest complexes was obtained by 1D and 2D NMR spectroscopy and molecular modelling. Our studies reveal a structure–binding property relationship for a series of structurally diverse aromatic alkaloids with the new receptor and higher binding affinity for the class of harmala alkaloids. To our knowledge, this is the first example of a chromophoric macrocyclic host employed as a molecular probe for the recognition of aromatic alkaloids.     

Exploring Multi-Subsite Binding Pockets in Proteins: DEEP-STD NMR Fingerprinting and Molecular Dynamics Unveil a Cryptic Subsite at the GM1 Binding Pocket of Cholera Toxin B

Ligand-based NMR techniques to study protein–ligand interactions are potent tools in drug design. Saturation transfer difference (STD) NMR spectroscopy stands out as one of the most versatile techniques, allowing screening of fragments libraries and providing structural information on binding modes. Recently, it has been shown that a multi-frequency STD NMR approach, differential epitope mapping (DEEP)-STD NMR, can provide additional information on the orientation of small ligands within the binding pocket. Here, the approach is extended to a so-called DEEP-STD NMR fingerprinting technique to explore the binding subsites of cholera toxin subunit B (CTB). To that aim, the synthesis of a set of new ligands is presented, which have been subject to a thorough study of their interactions with CTB by weak affinity chromatography (WAC) and NMR spectroscopy. Remarkably, the combination of DEEP-STD NMR fingerprinting and Hamiltonian replica exchange molecular dynamics has proved to be an excellent approach to explore the geometry, flexibility, and ligand occupancy of multi-subsite binding pockets. In the particular case of CTB, it allowed the existence of a hitherto unknown binding subsite adjacent to the GM1 binding pocket to be revealed, paving the way to the design of novel leads for inhibition of this relevant toxin.     

Analysis of Ah receptor binding affinities of polybrominated diphenyl ethers via in silico molecular docking and 3D-QSAR

Polybrominated diphenyl ethers (PBDEs) have become ubiquitous contaminations due to their use as flame retardants. The structural similarity of PBDE to some dioxin-like compounds suggested that they may share similar toxicological effects: they might activate the aryl hydrocarbon receptor (AhR) signal transduction pathway and thus might have adverse effects on wildlife and humans. In this study, in silico computational workflow combining molecular docking and three-dimensional quantitative structure–activity relationship (3D-QSAR) was performed to investigate the binding interactions between PBDEs and AhR and the structural features affecting the AhR binding affinity of PBDE. The molecular docking showed that hydrogen-bond and hydrophobic interactions were the major driving forces for the binding of ligands to AhR, and several key amino acid residues were also identified. The CoMSIA model was developed from the conformations obtained from molecular docking and exhibited satisfactory results as q ² of 0.605 and r ² of 0.996. Furthermore, the derived model had good robustness and statistical significance in both internal and external validations. The 3D contour maps generated from CoMSIA provided important structural features influence the binding affinity. The obtained results were beneficial to better understand the toxicological mechanism of PBDEs.     

Nucleotide recognition in water by a guanidinium-based artificial tweezer receptor

The water-soluble tweezer receptor 1 with two symmetric peptidic arms, which are connected by an aromatic scaffold and contain lysine, phenylalanine, and a guanidinium-based anion-binding site as headgroup, has been synthesized. UV/Vis-derived Job plots show that the receptor forms 1:1 complexes with nucleotides and phosphate in buffered water at neutral pH. Binding constants have been determined by fluorescence and UV/Vis spectroscopy. All nucleotides tested were bound very efficiently, even in pure water, with binding constants between 10(4) and 10(5) M(-1) . Interestingly, all mononucleotides were bound much stronger than phosphate by a factor of at least 5 to 10. Furthermore 1 favors the binding of adenosine monophosphate (AMP) over adenosine diphosphate (ADP) and adenosine triphosphate (ATP), which is unprecedented for artificial nucleotide receptors reported so far. According to NMR spectroscopy and molecular modeling studies, the efficient binding is a result of strong electrostatic contacts supported by π-π interactions with the nucleobase within the cavity-shaped receptor.     

Supramolecular Recognition of Amino Acids by Twisted Cucurbit[14]uril

Binding interactions between twisted cucurbit[14]uril (tQ[14]) and twenty standard amino acids (AAs) have been investigated by NMR spectroscopy and isothermal titration calorimetry (ITC) in aqueous HCl solutions and in DMSO. The results showed that tQ[14] displays clear binding affinity for AAs with a positively charged side chain or containing an aromatic ring, but weaker binding affinity for AAs with hydrophobic or polar side chains, with the binding mode depending on the type of side chain present in the AAs.     

不同功能基团的有机盐作为低相对分子质量凝胶因子对原油的成胶性能

低相对分子质量凝胶作为治理原油泄漏的材料具有一些优异的性能,但其在水油相中的成胶规律仍不明确,本文以有机酸及有机胺为原料设计合成了一系列有机盐。 通过核磁共振波谱仪(NMR)、傅里叶红外光谱仪(FTIR)、扫描电子显微镜(SEM)和倒挂实验等技术手段表征了有机盐的结构和成胶性能。 结果表明,有机盐A3C3(A3:肉桂酸;C3:月桂胺)在室温下能使原油形成稳定凝胶,成胶溶度为质量分数6%,该分子在溶剂中自组装形成树枝状三维网状结构。 凝胶的形成需要分子间π-π堆积作用、氢键、静电作用和范德华力等多种作用力协同作用,分子间的π-π堆积作用有利于凝胶的形成,含有多个苯环的凝胶因子更易在芳香族溶剂中形成凝胶。 这些成胶规律对处理原油泄漏的低相对分子质量凝胶因子的设计合成具有实际的指导意义。     

Selective recognition of aromatic hydrocarbons by endo-functionalized molecular tubes via C/N-H…π interactions

Aromatic hydrocarbons can be selectively recognized by four endo-functionalized molecular tubes through C/N-H...π interactions in nonpolar media with binding constants up to 1580 L/mol.