Molecular Recognition Studies Research Articles

Chiral Recognition on Bare Gold Surfaces by Quartz Crystal Microbalance.

Quartz crystal microbalance (QCM) is one of the powerful tools for the studies of molecular recognition and chiral discrimination. Its efficiency mainly relies on the design of the functional sensitive layer on the electrode surface. However, the organic sensitive layer may easily cause dissipation of oscillation or detachment and weaken the signal transfer during the molecular recognition processes. In this work, we reveal for the first time that the bare metal surface without the organic selector layer has the capability for chiral recognition in the QCM system. During the adsorption of various chiral amino acids, relatively higher selectivity of D-enantiomers on gold (Au) surface was shown by the QCM detection. Based on analyses of the surface crystalline structure and density functional theory calculations, we demonstrate that the chiral nature of Au surface plays an important role in the selective binding of specific D-amino acids. These results may open new insights on chiral detection by QCM system. It will also promote the construction of novel chiral sensing systems with both efficient detection and separation capability.

Chiral Recognition on Bare Gold Surfaces by Quartz Crystal Microbalance

AbstractQuartz crystal microbalance (QCM) is one of the powerful tools for the studies of molecular recognition and chiral discrimination. Its efficiency mainly relies on the design of the functional sensitive layer on the electrode surface. However, the organic sensitive layer may easily cause dissipation of oscillation or detachment and weaken the signal transfer during the molecular recognition processes. In this work, we reveal for the first time that the bare metal surface without the organic selector layer has the capability for chiral recognition in the QCM system. During the adsorption of various chiral amino acids, relatively higher selectivity of D‐enantiomers on gold (Au) surface was shown by the QCM detection. Based on analyses of the surface crystalline structure and density functional theory calculations, we demonstrate that the chiral nature of Au surface plays an important role in the selective binding of specific D‐amino acids. These results may open new insights on chiral detection by QCM system. It will also promote the construction of novel chiral sensing systems with both efficient detection and separation capability.

Anti-Metatype Antibody Screening, Sandwich Immunoassay Development, and Structural Insights for β-Lactams Based on Penicillin Binding Protein.

Theoretically, sandwich immunoassay is more sensitive and has a wider working range than that of competitive format. However, it has been thought that small molecules cannot be detected by the sandwich format due to their limited size. In the present study, we proposed a novel strategy for achieving sandwich immunoassay of β-lactams with low molecular weights. Firstly, five β-lactam antibiotics were selected to bind with penicillin binding protein (PBP)2x* to form complexes. Then, monoclonal and polyclonal antibodies against PBP2x*-β-lactams complexes were produced by animal immunization. Subsequently, the optimal pairing antibodies were utilized to establish sandwich immunoassay for detection of 18 PBP2x*-β-lactam complexes. Among them, ceftriaxone could be detected at as low as 1.65 ng/mL with working range of 1–1000 ng/mL in milk. To reveal the detection mechanism, computational chemistry and molecular recognition study were carried out. The results showed that β-lactams with a large size and complex structures maybe conducive to induce conformational changes of PBP2x*, and then exhibit greater possibility of being detected by sandwich immunoassay after combination with PBP2x*. This study provides insights for subsequent investigations of anti-metatype antibody screening and sandwich immunoassay establishment for small-molecule detection.

Shedding Light on the Molecular Recognition of Sub-Kilodalton Macrocyclic Peptides on Thrombin by Supervised Molecular Dynamics.

Macrocycles are attractive structures for drug development due to their favorable structural features, potential in binding to targets with flat featureless surfaces, and their ability to disrupt protein–protein interactions. Moreover, large novel highly diverse libraries of low-molecular-weight macrocycles with therapeutically favorable characteristics have been recently established. Considering the mentioned facts, having a validated, fast, and accurate computational protocol for studying the molecular recognition and binding mode of this interesting new class of macrocyclic peptides deemed to be helpful as well as insightful in the quest of accelerating drug discovery. To that end, the ability of the in-house supervised molecular dynamics protocol called SuMD in the reproduction of the X-ray crystallography final binding state of a macrocyclic non-canonical tetrapeptide—from a novel library of 8,988 sub-kilodalton macrocyclic peptides—in the thrombin active site was successfully validated. A comparable binding mode with the minimum root-mean-square deviation (RMSD) of 1.4 Å at simulation time point 71.6 ns was achieved. This method validation study extended the application domain of the SuMD sampling method for computationally cheap, fast but accurate, and insightful macrocycle–protein molecular recognition studies.

Hydrophobic Moiety of Capsaicinoids Haptens Enhancing Antibody Performance in Immunoassay: Evidence from Computational Chemistry and Molecular Recognition.

We previously found that the immune response to haptens is positively correlated with molecular hydrophobicity. The antibodies used in immunoassays for capsaicinoids (CPCs) in waste oil suffer from low affinity and loose recognition to structural analogues. To address this issue, four new haptens (hapten1-4), maximally exposing the hydrophobic alkane chain (noncommon moiety of CPCs), were designed and expected to produce antibodies with high affinity and accurate recognition to CPCs based upon our findings. The assumption was first evidenced by computational chemistry and animal immunization successively. Compared with four reported haptens (hapten5-8) that expose the hydrophilic vanillyl amide moiety (common structure of CPCs and other vanillin alkaloids), antisera from hapten1-4 showed an approximately 1000-fold increase in affinity and significantly improved recognition profiles for CPCs. The molecular recognition study showed that the high affinity of the antibody from new haptens mainly originated from hydrophobic forces. An indirect competitive enzyme-linked immunosorbent assay based on a monoclonal antibody from hapten1 was developed and exhibited limits of detection as low as 0.73-3.29 μg/kg for four CPCs in oils and with insignificant cross-reactivities for other eight vanillin alkaloids, which have been never achieved in previous reports.

A Comprehensive Review of Cholinesterase Modeling and Simulation.

The present article reviews published efforts to study acetylcholinesterase and butyrylcholinesterase structure and function using computer-based modeling and simulation techniques. Structures and models of both enzymes from various organisms, including rays, mice, and humans, are discussed to highlight key structural similarities in the active site gorges of the two enzymes, such as flexibility, binding site location, and function, as well as differences, such as gorge volume and binding site residue composition. Catalytic studies are also described, with an emphasis on the mechanism of acetylcholine hydrolysis by each enzyme and novel mutants that increase catalytic efficiency. The inhibitory activities of myriad compounds have been computationally assessed, primarily through Monte Carlo-based docking calculations and molecular dynamics simulations. Pharmaceutical compounds examined herein include FDA-approved therapeutics and their derivatives, as well as several other prescription drug derivatives. Cholinesterase interactions with both narcotics and organophosphate compounds are discussed, with the latter focusing primarily on molecular recognition studies of potential therapeutic value and on improving our understanding of the reactivation of cholinesterases that are bound to toxins. This review also explores the inhibitory properties of several other organic and biological moieties, as well as advancements in virtual screening methodologies with respect to these enzymes.

Host assisted molecular recognition by human serum albumin: Study of molecular recognition controlled protein/drug mimic binding in a microfluidic channel

Human serum albumin (HSA) plays a pivotal role in drug release from its delivery vehicles such as cyclodextrins (CDs) by binding to the drugs. Here molecular recognition and binding of a drug mimic (CD1) to HSA have been explored in a microfluidic channel when CD1 is encapsulated in β-cyclodextrin (βCD) and heptakis(2,3,6-tri-O-methyl)-β-cyclodextrin (TRIMEB), respectively, to investigate whether change of the host vehicle modulate the rate of drug binding to the serum protein. Molecular recognition of βCD encapsulated CD1 by HSA occurs by the conformational selection fit mechanism leading to rapid binding of CD1 to HSA (k1 ~ 700 s‐11) when the βCD/CD1 complex interacts with HSA. In contrary, HSA recognizes CD1 encapsulated in TRIMEB by an induced fit mechanism leading to a significantly slower binding rate (k1 ~ 20.8 s−1) of the drug mimic to the protein. Thus molecular recognition controls the rate of HSA binding by CD1 which in turn modulates the rate of delivery of the drug mimic from its macrocyclic hosts. The remarkable change in the molecular recognition pathway of CD1 by HSA, upon change of the host from βCD to TRIMEB, originates from significantly different conformational flexibility of the host/drug mimic complexes.

FMODB: The World's First Database of Quantum Mechanical Calculations for Biomacromolecules Based on the Fragment Molecular Orbital Method.

We developed the world's first web-based public database for the storage, management, and sharing of fragment molecular orbital (FMO) calculation data sets describing the complex interactions between biomacromolecules, named FMO Database (https://drugdesign.riken.jp/FMODB/). Each entry in the database contains relevant background information on how the data was compiled as well as the total energy of each molecular system and interfragment interaction energy (IFIE) and pair interaction energy decomposition analysis (PIEDA) values. Currently, the database contains more than 13 600 FMO calculation data sets, and a comprehensive search function implemented at the front-end. The procedure for selecting target proteins, preprocessing the experimental structures, construction of the database, and details of the database front-end were described. Then, we demonstrated a use of the FMODB by comparing IFIE value distributions of hydrogen bond, ion-pair, and XH/π interactions obtained by FMO method to those by molecular mechanics approach. From the comparison, the statistical analysis of the data provided standard reference values for the three types of interactions that will be useful for determining whether each interaction in a given system is relatively strong or weak compared to the interactions contained within the data in the FMODB. In the final part, we demonstrate the use of the database to examine the contribution of halogen atoms to the binding affinity between human cathepsin L and its inhibitors. We found that the electrostatic term derived by PIEDA greatly correlated with the binding affinities of the halogen containing cathepsin L inhibitors, indicating the importance of QM calculation for quantitative analysis of halogen interactions. Thus, the FMO calculation data in FMODB will be useful for conducting statistical analyses to drug discovery, for conducting molecular recognition studies in structural biology, and for other studies involving quantum mechanics-based interactions.

Molecular Recognition in Water Using Macrocyclic Synthetic Receptors.

Molecular recognition in water using macrocyclic synthetic receptors constitutes a vibrant and timely research area of supramolecular chemistry. Pioneering examples on the topic date back to the 1980s. The investigated model systems and the results derived from them are key for furthering our understanding of the remarkable properties exhibited by proteins: high binding affinity, superior binding selectivity, and extreme catalytic performance. Dissecting the different effects contributing to the proteins' properties is severely limited owing to its complex nature. Molecular recognition in water is also involved in other appreciated areas such as self-assembly, drug discovery, and supramolecular catalysis. The development of all these research areas entails a deep understanding of the molecular recognition events occurring in aqueous media. In this review, we cover the past three decades of molecular recognition studies of neutral and charged, polar and nonpolar organic substrates and ions using selected artificial receptors soluble in water. We briefly discuss the intermolecular forces involved in the reversible binding of the substrates, as well as the hydrophobic and Hofmeister effects operating in aqueous solution. We examine, from an interdisciplinary perspective, the design and development of effective water-soluble synthetic receptors based on cyclic, oligo-cyclic, and concave-shaped architectures. We also include selected examples of self-assembled water-soluble synthetic receptors. The catalytic performance of some of the presented receptors is also described. The latter process also deals with molecular recognition and energetic stabilization, but instead of binding ground-state species, the targets become elusive counterparts: transition states and other high-energy intermediates.

An intramolecularly self-templated synthesis of macrocycles: self-filling effects on the formation of prismarenes.

Ethyl- and propyl-prism[6]arenes are obtained in high yields and in short reaction times, independent of the nature and size of the solvent, in the cyclization of 2,6-dialkoxynaphthalene with paraformaldehyde. PrS[6]Et or PrS[6]nPr adopt, both in solution and in the solid state, a folded cuboid-shaped conformation, in which four inward oriented alkyl chains fill the cavity of the macrocycle. On these bases, we proposed that the cyclization of PrS[6]Et or PrS[6]nPr occurs through an intramolecular thermodynamic self-templating effect. In other words, the self-filling of the internal cavity of PrS[6]Et or PrS[6]nPr stabilizes their cuboid structure, driving the equilibrium toward their formation. Molecular recognition studies, both in solution and in the solid state, show that the introduction of guests into the macrocycle cavity forces the cuboid scaffold to open, through an induced-fit mechanism. An analogous conformational change from a closed to an open state occurs during the endo-cavity complexation process of the pentamer, PrS[5]. These results represent a rare example of a thermodynamically controlled cyclization process driven through an intramolecular self-template effect, which could be exploited in the synthesis of novel macrocycles.

AquaPELE: A Monte Carlo-Based Algorithm to Sample the Effects of Buried Water Molecules in Proteins

Water is frequently found inside proteins, carrying out important roles in catalytic reactions or molecular recognition tasks. Therefore, computational models that aim to study protein-ligand interactions usually have to include water effects through explicit or implicit approaches to obtain reliable results. While full explicit models might be too computationally daunting for some applications, implicit models are normally faster but omit some of the most important contributions of water. This is the case of our in-house software, called protein energy landscape exploration (PELE), which uses implicit models to speed up conformational explorations as much as possible; the lack of explicit water sampling, however, limits its model. In this work, we confront this problem with the development of aquaPELE. It is a new algorithm that extends the exploration capabilities while keeping efficiency as it employs a mixed implicit/explicit approach to also take into account the effects of buried water molecules. With an additional Monte Carlo (MC) routine, a set of explicit water molecules is perturbed inside protein cavities and their effects are dynamically adjusted to the current state of the system. As a result, this implementation can be used to predict the principal hydration sites or the rearrangement and displacement of conserved water molecules upon the binding of a ligand. We benchmarked this new tool focusing on estimating ligand binding modes and hydration sites in cavities with important interfacial water molecules, according to crystallographic structures. Results suggest that aquaPELE sets a fast and reliable alternative for molecular recognition studies in systems with a strong water-dependency.

Effect of Substituent of Roof Shape Amines on the Molecular Recognition of Optically Active Acids by NMR Spectroscopy

AbstractOptically active roof shape derivatives of benzyl amines were synthesized and screened as chiral solvating agents for the study of molecular recognition of chiral compounds by NMR and UV spectroscopy. The effect of substituents on the aromatic ring of benzyl unit is investigated to assess the binding ability with the analyte and an experimental correlation is observed. Altering the substituents on the ring of benzyl amine and the acidic substrates, significantly influence the CSA interactions. The supramolecular interactions between acid and amine in the single crystal X‐ray analysis of the diastereomeric salts provide further insight of the mode of molecular recognition.

Direct attachment of suspension cells to PDA surface and its application in suspension-cell QCM biosensor

Mussel-inspired polydopamine (PDA) has been widely applied for functionalizing various material surfaces through a simple and versatile approach. Several studies have demonstrated the adhesion-promotion properties of the PDA surface for adherent cells. However, few reports are associated with suspension cells. In this study, the attachment property of suspension cells on the PDA surface was investigated for the first time, where we found that the suspension cells can be directly attached to the PDA coating without any secondary modifications. To exemplify the potential of this property, a PDA-based suspension-cell QCM biosensor was fabricated. The biosensor showed a high degree of repeatability and stability as well as low nonspecific binding to the irrelevant protein. The real-time and label-free evaluation of the binding of a diverse range of lectins on three different types of suspension cells indicated the variation of the glycosylation on these cell surfaces. Furthermore, the kinetic evaluation of the protein-carbohydrate interactions on the suspension cell surface was also performed. This work reports a new strategy for fabrication of PDA-based suspension-cell QCM biosensor via simple and efficient immobilization of suspension cells, which provides a simple way for the study of the complex molecular recognition on suspension cell surfaces.

Electrochemical investigation of the intermolecular hydrogen bond on the surface of self-assembled monolayer

Intermolecular hydrogen bond, as a kind of weak interaction, plays a fundamental and important role in the molecular recognition process on cell membrane surface. Based on the complexity of the cell membrane and its microenvironment, the self-assembled monolayer with clear structure is used to simulate the cell membrane to construct the simulation system. The intermolecular hydrogen bond on the surface of the self-assembled is explored at the molecular level, which provided a new perspective for the study of molecular recognition on the cell membrane surface. In this paper, combined with the advantages of scanning electrochemical microscopy, the progress on the intermolecular hydrogen bond on the surface of self-assembled monolayer is introduced from the electrochemical viewpoint.

Synthesis, Molecular Recognition Study and Liquid Membrane-Based Applications of Highly Lipophilic Enantiopure Acridino-Crown Ethers.

New highly lipophilic enantiopure crown ethers containing a heterocyclic unit have been synthesized. Phase transport, UV-Vis- and fluorescence spectrophotometric investigations as well as electrochemical studies on the complexation of the new macrocycles with several amine and amino acid derivatives were also carried out. Achiral amines were used for studying the structural preference of the new macrocycles. Among the studied structural features of the guest molecules, the intermolecular π-π interaction showed the most significant effect on complexation, which made the aralkylamine-type compounds the most preferable guest molecules. The studied liquid membrane-based applications and photophysical investigations showed appreciable enantiomeric recognition toward some aralkylamine model compounds with homochiral preferences. New crown ether derivatives (R,R)-2 and (S,S)-2 were successfully applied as enantioselective carrier and sensor molecules.

No, Not That Way, the Other Way: Creating Active Sites in Self-Assembled Host Molecules

This Account describes our efforts over the last decade to synthesize self-assembled metal–ligand cage complexes that display reactive functional groups on their interiors. This journey has taken us down a variety of research avenues, including studying the mechanism of reversible self-assembly, analyzing ligand self-sorting properties, post-assembly reactivity, molecular recognition, and binding studies, and finally reactivity and catalysis. Each of these individual topics are discussed here, as are the lessons learned along the way and the future research outlook. These self-assembled hosts are the closest mimics of enzymes to date, as they are capable of size- and shape-selective molecular recognition, substrate activation and turnover, as well as showing less common ‘biomimetic’ properties such as the ability to employ cofactors in reactivity, and alter the prevailing mechanism of the catalyzed reactions.1 Introduction2 Paddlewheels and Self-Sorting Behavior3 First-Row Transition-Metal-Mediated Assembly: Sorting and Stereochemical Control4 Post-Assembly Reactivity5 Molecular Recognition and Catalysis6 Conclusions and Outlook

Shape-Induced Selective Separation of Ortho-substituted Benzene Isomers Enabled by Cucurbit[7]uril Host Macrocycles

Summary The separation of benzene derivatives is energy intensive and laborious as a result of the overlapping physicochemical properties of these isomers. Here, we report on the separation of ortho-disubstituted benzene isomers using cucurbit[7]uril (CB7) aqueous solution with more than 92% selectivity. Thermodynamic and kinetic analysis proves that the ortho-isomer has stronger binding ability and slower decomplexation rate constant than the para- and meta-isomers when hosted by CB7. Optimized host-guest models indicate that the ortho-isomer with the smallest aspect ratio well matches the spherical interior cavity of CB7, resulting in highly stable complexes. Furthermore, laboratory scale-up experiments using commercial xylenes and C8 aromatic fraction of pyrolysis gasoline proved that CB7 is able to separate ortho-xylene (OX) with a remarkable selectivity of up to 83%. We believe that this work accentuates the role of molecular recognition studies using macrocyclic hosts to improve the quality and energy bill of critical industrial separations. Video Abstract Download : Download video (27MB)

Fluorinated carbohydrates as chemical probes for molecular recognition studies. Current status and perspectives.

This review provides an extensive summary of the effects of carbohydrate fluorination with regard to changes in physical, chemical and biological properties with respect to regular saccharides. The specific structural, conformational, stability, reactivity and interaction features of fluorinated sugars are described, as well as their applications as probes and in chemical biology.

A top-down chemo-enzymatic approach towards N-acetylglucosamine-N-acetylmuramic oligosaccharides: Chitosan as a reliable template

An unprecedented approach towards oligosaccharides containing N-acetylglucosamine-N-acetylmuramic (NAG-NAM) units was developed. These novel bacterial cell wall surrogates were obtained from chitosan via a top down approach involving both chemical and enzymatic reactions. The chemical modification of chitosan using a molecular clamp based strategy, allowed obtaining N-acetylglucosamine-N-acetylmuramic (NAG-NAM) containing oligomers. Intercalation of NAM residues was confirmed through the analysis of oligosaccharide fragments from enzymatic digestion and it was found that this route affords NAG-NAM containing oligosaccharides in 33% yield. These oligosaccharides mimic the carbohydrate basic skeleton of most bacterial cell surfaces. The oligosaccharides prepared are biologically relevant and will serve as a platform for further molecular recognition studies with different receptors and enzymes of both bacterial cell wall and innate immune system. This strategy combining both chemical modification and enzymatic digestion provides a novel and simple route for an easy access to bacterial cell wall fragments – biologically important targets.

Crown Ether Ditopic Receptors for Ammonium Salts with High Affinity for Amino Acid Ester Salts

Two bis crown ether receptors were synthesized and tested as host molecules for protonated forms of alkyl amines and amino acid esters. Molecular recognition studies were conducted in CH2Cl2:MeOH (92:8) by spectrophotometric UV/Vis titrations andby spectrometric 1H NMR titrations in CDCl3. The calculated binding constants are in the range 102-105 M-1. A high affinity for L-amino acid methyl ester derivatives was found. A theoretical study at the DFT level of the synthesized receptor and some analog ligands with three different ammonium ions helps to rationalize the experimentally found trends.