Showing papers in "Journal of Molecular Recognition in 2014"
TL;DR: A survey of the literature covering the development of molecular imprinting science and technology over the years 2004–2011 and efforts to apply these polymeric materials to a range of application areas is presented.
Abstract: Herein, we present a survey of the literature covering the development of molecular imprinting science and technology over the years 2004-2011. In total, 3779 references to the original papers, rev ...
413 citations
TL;DR: It was found that each ABR differs significantly in its amino acid composition and tends to bind different types of amino acids at the surface of proteins, and the combined preference of the six ABRs does not allow epitopes to be distinguished from the rest of the protein surface.
Abstract: The concept of antibody specificity is analyzed and shown to reside in the ability of an antibody to discriminate between two antigens. Initially antibody specificity was attributed to sequence differences in complementarity-determining regions (CDRs) but as increasing numbers of crystallographic antibody-antigen complexes were elucidated, specificity was analyzed in terms of six antigen-binding regions (ABRs) that only roughly correspond to CDRs. It was found that each ABR differs significantly in its amino acid composition and tends to bind different types of amino acids at the surface of proteins. In spite of these differences, the combined preference of the six ABRs does not allow epitopes to be distinguished from the rest of the protein surface. These findings explain the poor success of past and newly proposed methods for predicting protein epitopes. Antibody polyspecificity refers to the ability of one antibody to bind a large variety of epitopes in different antigens and this property explains how the immune system develops an antibody repertoire that is able to recognize every antigen the system is likely to encounter. Antibody heterospecificity arises when an antibody reacts better with another antigen than with the one used to raise the antibody. As a result an antibody may sometimes appear to have been elicited by an antigen with which it is unable to react. The implications of antibody polyspecificity and heterospecificity in vaccine development are pointed out.
103 citations
TL;DR: The determination of dissociation constant (Kd) values revealed that aptamers generated against aflatoxin M1 exhibited Kd values in the range of 35–1515 nM, and it is interesting that one aptamer with no conserved motif or G‐quadruplex had lowest Kd value (KD = 35 NM).
Abstract: In the present work, aptamers against aflatoxin M1 and aflatoxin B1 were generated and tested for creating proof of principle of recognition of aflatoxin M1 by generated aptamers. The aptamers were selected through the process referred as systematic evolution of ligands by exponential enrichment. A total of 41 different aptamer (36 aptamers for aflatoxin M1 and 5 for aflatoxin B1) sequences were obtained. The determination of dissociation constant (Kd ) values revealed that aptamers generated against aflatoxin M1 exhibited Kd values in the range of 35-1515 nM. Selected aptamers were grouped on the basis of the presence of common motifs or G-quadruplex. We find it interesting that one aptamer with no conserved motif or G-quadruplex had lowest Kd value (Kd = 35 nM). This structural motif is very distinct from motifs present in other aptamers. The Kd values of selected aptamers for aflatoxin B1 were in the range of 96-221 nM. One aptamer from each group was further tested for its ability to be used in aptasensor. The aptamer recognized aflatoxin M1 as indicated by color change (red to purple or blue) of aptamer-coated gold nanoparticles in the presence of 250-500 nM aflatoxin M1. The aptamers can be used in developing methods for detection/estimation/separation of aflatoxin or antidote for aflatoxin toxicity.
70 citations
TL;DR: The sol–gel MIP has a high specificity and capacity for the drug in ethanol, a solvent compatible with drug formulation and biomedical applications, and the results were modelled by Fick's law of diffusion and the power law.
Abstract: Molecularly imprinted polymers (MIPs) for salicylic acid were synthesized and evaluated in aqueous environments in the aim to apply them as drug delivery carriers. One organic MIP and one inorganic MIP based on the sol-gel process were synthesized. The organic MIP was prepared by radical polymerization using the stoichiometric functional monomer, 1-(4-vinylphenyl)-3-(3,5-bis(trifluoromethyl)phenyl)urea, which can establish strong electrostatic interactions with the -COOH of salicylic acid. The sol-gel MIP was prepared with 3-(aminopropyl)triethoxysilane and trimethoxyphenylsilane, as functional monomers and tetraethyl orthosilicate as the crosslinker. While the organic MIPs bound the target specifically in acetonitrile, they exhibited lower binding in the presence of water, although the imprinting factor increased under these conditions, due to reduced non-specific binding. The sol-gel MIP has a high specificity and capacity for the drug in ethanol, a solvent compatible with drug formulation and biomedical applications. In vitro release profiles of the polymers in water were evaluated, and the results were modelled by Fick's law of diffusion and the power law. Analysis shows that the release mechanism was predominantly diffusion-controlled.
49 citations
TL;DR: The overall data supports the use of CN derivatives against quorum sensing‐mediated infections caused by S. pyogenes and confirms that speB is controlled by the regulation of luxS.
Abstract: The LuxS-based signalling pathway has an important role in physiological and pathogenic functions that are capable of causing different infections. In the present study, cinnamaldehyde (CN) and their derivatives were evaluated for their inhibitory efficiency against LuxS by molecular modelling, docking, dynamics and free-energy calculations. Sequence and structure-similarity analysis of LuxS protein, five different amino acids were found to be highly conserved, of which GLY128 was identified as the key residue involved in the effective binding of the ligands. Quantum-polarized ligand docking protocol showed that 2nitro and 4nitro CN has a higher binding efficiency than CN, which very well corroborates with the in vitro studies. COMSTAT analysis for the microscopic images of the S. pyogenes biofilm showed that the ligands have antibiofilm potential. In addition, the results of quantitative polymerase chain reaction (qPCR) analysis revealed that the transcripts treated with the compounds showed decrease in luxS expression, which directly reflects with the reduction in expression of speB. No substantial effect was observed on the virulence regulator (srv) transcript. These results confirm that speB is controlled by the regulation of luxS. The decreased rate of S. pyogenes survival in the presence of these ligands envisaged the fact that the compounds could readily enhance opsonophagocytosis with the reduction of virulence factor secretion. Thus, the overall data supports the use of CN derivatives against quorum sensing-mediated infections caused by S. pyogenes.
44 citations
TL;DR: Thiophene‐based diimine and monoimine were synthesized in a single step and their cation binding affinity was tested using colorimetric and UV–vis spectral studies, and R2 was also successfully demonstrated as a fluorescent probe for detecting Cu2+ ions in living cells.
Abstract: Thiophene-based diimine (R1) and monoimine (R2) were synthesized in a single step, and their cation binding affinity was tested using colorimetric and UV–vis spectral studies. R1 selectively shows a colorimetric turn-on response for Pb2+, Hg2+ ions and colorimetric turn-off with Sn2+ ions, and R2 shows visual response for Cu2+ and Hg2+ over other examined metal ions in aqueous medium. R1 forms 1:1 complex with Pb2+, Hg2+, and Sn2+ and exhibits fluorescence quenching, whereas R2 shows 2:1 complex with Hg2+, Cu2+ and shows fluorescence enhancement. The structural and electronic properties of the sensors and their metal complexes were also investigated using Density Functional Theory calculations. R2 was also successfully demonstrated as a fluorescent probe for detecting Cu2+ ions in living cells. Copyright © 2014 John Wiley & Sons, Ltd.
44 citations
TL;DR: First insight is provided into MDP recognition and CARD–CARD interaction in the zebrafish that will be useful to understand the molecular basis of NOD signaling in a broader perspective.
Abstract: Nucleotide binding and oligomerization domain (NOD2) is a key component of innate immunity that is highly specific for muramyl dipeptide (MDP)—a peptidoglycan component of bacterial cell wall MDP recognition by NOD2–leucine rich repeat (LRR) domain activates NF-κB signaling through a protein–protein interaction between caspase activating and recruitment domains (CARDs) of NOD2 and downstream receptor interacting and activating protein kinase 2 (RIP2) Due to the lack of crystal/NMR structures, MDP recognition and CARD–CARD interaction are poorly understood Herein, we have predicted the probable MDP and CARD–CARD binding surfaces in zebrafish NOD2 (zNOD2) using various in silico methodologies The results show that the conserved residues Phe819, Phe871, Trp875, Trp929, Trp899, and Arg845 located at the concave face of zNOD2–LRR confer MDP recognition by hydrophobic and hydrogen bond (H-bond) interactions Molecular dynamics simulations reveal a stable association between the electropositive surface on zNOD2–CARDa and the electronegative surface on zRIP2–CARD reinforced mostly by H-bonds and electrostatic interactions Importantly, a 35 A salt bridge is observed between Arg60 of zNOD2–CARDa and Asp494 of zRIP2–CARD Arg11 and Lys53 of zNOD2–CARDa and Ser498 and Glu508 of zRIP2–CARD are critical residues for CARD–CARD interaction and NOD2 signaling The 27 A H-bond between Lys104 of the linker and Glu508 of zRIP2–CARD suggests a possible role of the linker for shaping CARD–CARD interaction These findings are consistent with existing mutagenesis data We provide first insight into MDP recognition and CARD–CARD interaction in the zebrafish that will be useful to understand the molecular basis of NOD signaling in a broader perspective Copyright © 2014 John Wiley & Sons, Ltd
36 citations
TL;DR: A molecularly imprinted composite cryogel exhibited a high binding capacity and selectivity for hemoglobin in the presence of immunoglobulin G, albumin and myoglobin and can be reused many times with no apparent decrease in hemoglobin adsorption capacity.
Abstract: A molecularly imprinted composite cryogel (MICC) was prepared for depletion of hemoglobin from human blood prior to use in proteome applications. Poly(hydroxyethyl methacrylate) based MICC was prepared with high gel fraction yields up to 90%, and characterized by Fourier transform infrared spectrophotometer, scanning electron microscopy, swelling studies, flow dynamics and surface area measurements. MICC exhibited a high binding capacity and selectivity for hemoglobin in the presence of immunoglobulin G, albumin and myoglobin. MICC column was successfully applied in fast protein liquid chromatography system for selective depletion of hemoglobin for human blood. The depletion ratio was highly increased by embedding microspheres into the cryogel (93.2%). Finally, MICC can be reused many times with no apparent decrease in hemoglobin adsorption capacity. Copyright © 2014 John Wiley & Sons, Ltd.
34 citations
TL;DR: The results indicate that the novel nanocomposite show a high affinity for these heavy metals due to the presence of several good extractive sites, which are introduced to the synthesized nanocompositionite.
Abstract: Poly 1,8-diaminonaphthalene/multiwalled carbon nanotubes-COOH hybrid material as an effective sorbents in solid phase extraction has been developed for the separation and preconcentration of Cd(II) and Pb(II) at trace levels in environmental water samples. The results indicate that the novel nanocomposite show a high affinity for these heavy metals due to the presence of several good extractive sites, which are introduced to the synthesized nanocomposite The maximum adsorption capacity of the synthesized sorbent for cadmium and lead ions was found to be 101.2 and 175.2 mg g(-1) , respectively. The detection limits of this method were 0.09 and 0.7 ng ml(-1) for Cd(II) and Pb(II), respectively.
34 citations
TL;DR: It is reported in this paper that the binding of coumarin 6 (C6) to DNA can be tuned by complexing it with host structures, viz.
Abstract: We report in this paper that the binding of coumarin 6 (C6) to DNA can be tuned by complexing it with host structures, viz. β-cyclodextrin (β-CD) and C-hexylpyrogallol-4-arene (C-HPA). Because host molecules are used as carriers of small molecules onto target sites, the exposed part of the guest molecule needs to be found out, and the relationship between the host : guest ratio and the mode of binding with the target macromolecule, that is, the DNA needs to be analyzed, in order to comprehend the preferred binding moiety and tune the binding. In this paper, the formation of the inclusion complex of C6 with β-CD and with C-HPA is studied by UV-visible, fluorescence, 2D rotating-frame nuclear Overhauser effect correlation spectroscopy and diffusion-ordered spectroscopy nuclear magnetic resonance spectra and molecular modeling. C6 forms a 1:1 complex with β-CD and a 1:2 complex with C-HPA. The studies on the protonation of C6 in the presence and the absence of the host molecules suggest that the chromone part of C6 is outside the β-CD molecule, whereas it is fully covered by C-HPA. The binding of C6 with calf thymus DNA (ctDNA) occurs through intercalation and hydrogen bonding, and the host–guest structures remain intact on binding with ctDNA. The oxygens of the C6 molecules are exposed when inside the host molecules and aid in the hydrogen bonding with DNA. Copyright © 2014 John Wiley & Sons, Ltd.
33 citations
TL;DR: The mode of action of this stress‐related protein is demonstrated by performing comparative modeling of Q10RT8 and docking results indicated that Ser21, Ala22, Lys25, Asp68, Ala70, Glu73, and Arg74 are important determinant residues for functional interaction with the GTP ligand.
Abstract: In the present research, we have studied the inoculation effects of two root-associated plant growth-promoting rhizobacteria (PGPR) in rice and provide the pieces of evidence that the inoculation of the PGPR could potentially result in inducing the expression of the salt stress-related RAB18 plant gene under varying degrees of salinity stress. The sequenced putative gene of RAB18 of Oryza sativa in this study is 740 bp long, has a content of 44.4%, and a molecular weight of 492 102.00 Da. BLAST homology patterns revealed sequence similarity with the previously sequenced RAB in model plant species. We demonstrate the mode of action of this stress-related protein by performing comparative modeling of Q10RT8 (Os03g0146000 protein, homolog of the sequenced RAB18; O. sativa subsp. japonica) using energy minimization, molecular dynamic simulations, and molecular docking of a guanosine triphosphate (GTP) ligand with the protein. The docking results indicated that Ser21, Ala22, Lys25, Asp68, Ala70, Glu73, and Arg74 are important determinant residues for functional interaction with the GTP ligand. The present research contributes to the understanding of the PGPR inoculation in salinity stress. Additionally, it provides the layout of the understanding of the molecular interactions between RAB and GTP ligand. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: CCapto Adhere proved to offer a very strong binding of nucleic acids and could be used to isolate plasmid DNA from a crude Escherichia coli extract without protein and endotoxin contaminations.
Abstract: in Undetermined Multimodal chromatography is widely used for isolation of proteins because it often results in improved selectivity compared to conventional separation resins. The binding potential and chromatographic behavior of plasmid DNA have here been examined on a Capto Adhere resin. Capto Adhere is a recent multimodal chromatography material allowing molecular recognition between the ligand and target molecule, which is based on combined ionic and aromatic interactions. Capto Adhere proved to offer a very strong binding of nucleic acids. This property could be used to isolate plasmid DNA from a crude Escherichia coli extract. Using a stepwise NaCl gradient, pure plasmid DNA could be obtained without protein and endotoxin contaminations. The RNA fraction bound most strongly to the resin and could be eluted only at very high salt concentrations (2.0 M NaCl). The chromatographic separation behavior was very robust between pH values 6 and 9, and the dynamic binding capacity was estimated to 60 µg/ml resin. (Less)
TL;DR: The isothermal titration calorimetry technique supported by potentiometric titration data was used to study the interaction of zinc ions with pH buffer substances, namely 2‐(N‐morpholino)ethanesulfonic acid (Mes), piperazine‐N,N′‐bis(2‐ethanes sulfuronic acid) (Pipes), and dimethylarsenic acid (Caco).
Abstract: The isothermal titration calorimetry (ITC) technique supported by potentiometric titration data was used to study the interaction of zinc ions with pH buffer substances, namely 2-(N-morpholino)ethanesulfonic acid (Mes), piperazine-N,N'-bis(2-ethanesulfonic acid) (Pipes), and dimethylarsenic acid (Caco). The displacement ITC titration method with nitrilotriacetic acid as a strong, competitive ligand was applied to determine conditional-independent thermodynamic parameters for the binding of Zn(II) to Mes, Pipes, and Caco. Furthermore, the relationship between the proposed coordination mode of the buffers and the binding enthalpy has been discussed.
TL;DR: The molecular interaction mechanism between Fc fragment of IgG and a synthetic ligand (DAAG) was studied with molecular docking and dynamics simulation to shed light on the molecular mechanisms of DAAG for antibody purification.
Abstract: Affinity chromatography with synthetic ligands has been focused as the potential alternative to protein A-based chromatography for antibody capture because of its comparable selectivity and efficiency. Better understanding on the molecular interactions between synthetic ligand and antibody is crucial for improving and designing novel ligands. In this work, the molecular interaction mechanism between Fc fragment of IgG and a synthetic ligand (DAAG) was studied with molecular docking and dynamics simulation. The docking results on the consensus binding site (CBS) indicated that DAAG could bind to the CBS with the favorable orientation like a tripod for the top-ranked binding complexes. The ligand-Fc fragment complexes were then tested by molecular dynamics simulation at neutral condition (pH 7.0) for 10 ns. The results indicated that the binding of DAAG on the CBS of Fc fragment was achieved by the multimodal interactions, combining the hydrophobic interaction, electrostatic interaction, hydrogen bond, and so on. It was also found that multiple secondary interactions endowed DAAG with an excellent selectivity to Fc fragment. In addition, molecular dynamics simulation conducted at acidic condition (pH 3.0) showed that the departure of DAAG ligand from the surface of Fc fragment was the result of reduced interaction energies. The binding modes between DAAG and CBS not only shed light on the molecular mechanisms of DAAG for antibody purification but also provide useful information for the improvement of ligand design. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: The synthesis and evaluation of imprinted films for SPME by electropolymerisation of pyrrole alone or in the presence of ethylene glycol dimethacrylate is proposed and sulfadimethoxine, a sulfonamide antibiotic, was used as template molecule.
Abstract: Solid-phase microextraction (SPME) is widely used in analytical laboratories for the analysis of organic compounds, thanks to its simplicity and versatility. In the present work, the synthesis and evaluation of imprinted films for SPME by electropolymerisation of pyrrole alone or in the presence of ethylene glycol dimethacrylate is proposed. Sulfadimethoxine (SDM), a sulfonamide antibiotic, was used as template molecule. Initially, a molecularly imprinted polymer film was prepared by electropolymerisation of pyrrole onto a platinum foil, using SDM as template. The SDM template was removed by overoxidation. The behaviour of SDM on imprinted and non-imprinted polymers was investigated by differential pulse voltammetry, and a clear imprinting effect was observed, which was confirmed by rebinding experiments using both conventional and electrochemically enhanced-SPME. However, in general, the extraction efficiency was rather low (<6%) and unspecific interactions are too high. Attempts to increase extraction efficiency were unsuccessful, but the incorporation of ethylene glycol dimethacrylate to the films reduced unspecific interactions to a certain extent. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: Conductive composite films comprised of single‐walled carbon nanotubes coated with molecularly imprinted poly‐4‐vinylphenol are produced and characterized using ultraviolet and infrared spectroscopies, confirming the successful molecular imprinting of the film with cotinine.
Abstract: Conductive composite films comprised of single-walled carbon nanotubes coated with molecularly imprinted poly-4-vinylphenol are produced and characterized using ultraviolet and infrared spectroscopies, confirming the successful molecular imprinting of the film with cotinine. The electrical resistance of the imprinted film changes significantly upon binding cotinine, by more than 30 kΩ, while the unimprinted film in comparison elicits little response. Additionally, once the cotinine template desorbs from the film, the resistance of the imprinted film returns to a value close to the pre-adsorption baseline. Scanning electron microscopy is used to study the morphology of the film compared with the unimprinted control, and gas chromatography quantitatively confirms that the imprinted film selectively detects cotinine while discriminating against the structurally similar alkaloid, nicotine.
TL;DR: In this article, the development and findings of parameter-unlimited functional super-resolution microscopy are described, a technology based on the fluorescence imaging cycler (IC) principle capable of co-mapping thousands of distinct biomolecular assemblies at high spatial resolution and differentiation.
Abstract: Understanding biological systems at the level of their relational (emergent) molecular properties in functional protein networks relies on imaging methods, able to spatially resolve a tissue or a cell as a giant, non-random, topologically defined collection of interacting supermolecules executing myriads of subcellular mechanisms Here, the development and findings of parameter-unlimited functional super-resolution microscopy are described—a technology based on the fluorescence imaging cycler (IC) principle capable of co-mapping thousands of distinct biomolecular assemblies at high spatial resolution and differentiation (<40 nm distances) It is shown that the subcellular and transcellular features of such supermolecules can be described at the compositional and constitutional levels; that the spatial connection, relational stoichiometry, and topology of supermolecules generate hitherto unrecognized functional self-segmentation of biological tissues; that hierarchical features, common to thousands of simultaneously imaged supermolecules, can be identified; and how the resulting supramolecular order relates to spatial coding of cellular functionalities in biological systems A large body of observations with IC molecular systems microscopy collected over 20 years have disclosed principles governed by a law of supramolecular segregation of cellular functionalities This pervades phenomena, such as exceptional orderliness, functional selectivity, combinatorial and spatial periodicity, and hierarchical organization of large molecular systems, across all species investigated so far This insight is based on the high degree of specificity, selectivity, and sensitivity of molecular recognition processes for fluorescence imaging beyond the spectral resolution limit, using probe libraries controlled by ICs © 2013 The Authors Journal of Molecular Recognition published by John Wiley & Sons, Ltd
TL;DR: Some of the theoretical and experimental aspects of the affinity‐based methods are discussed and the protein consumption is evaluated to develop methods for the screening of further new compounds.
Abstract: This work describes the development of biophysical unbiased methods to study the interactions between new designed compounds and carbonic anhydrase II (CAII) enzyme. These methods have to permit both a screening of a series of sulfonamide derivatives and the identification of a lead compound after a thorough study of the most promising molecules. Interactions data were collected using surface plasmon resonance (SPR) and thermal shift assay (TSA). In the first step, experiments were performed with bovine CAII isoform and were extended to human CAII. Isothermal titration calorimetry (ITC) experiments were also conducted to obtain thermodynamics parameters necessary for the processing of the TSA data. Results obtained with this reference methodology demonstrate the effectiveness of SPR and TSA. KD values obtained from SPR data were in perfect accordance with ITC. For TSA, despite the fact that the absolute values of KD were quite different, the same affinity scale was obtained for all compounds. The binding affinities of the analytes studied vary by more than 50 orders of magnitude; for example, the KD value determined by SPR were 6 ± 4 and 299 ± 25 nM for compounds 1 and 3, respectively. This paper discusses some of the theoretical and experimental aspects of the affinity-based methods and evaluates the protein consumption to develop methods for the screening of further new compounds. The double interest of SPR, that is, for screening and for the quick thorough study of the interactions parameters (ka, kd, and KD), leads us to choose this methodology for the study of new potential inhibitors. Copyright © 2013 John Wiley & Sons, Ltd.
TL;DR: The binding pockets of about 4000 PDB protein‐ligand complexes were investigated and amino acid and interaction types were analyzed, showing that Gly, Glu, Arg, Asp, His, and Thr are more conserved than other amino acids.
Abstract: The large number of macromolecular structures deposited with the Protein Data Bank (PDB) describing complexes between proteins and either physiological compounds or synthetic drugs made it possible a systematic analysis of the interactions occurring between proteins and their ligands. In this work, the binding pockets of about 4000 PDB protein-ligand complexes were investigated and amino acid and interaction types were analyzed. The residues observed with lowest frequency in protein sequences, Trp, His, Met, Tyr, and Phe, turned out to be the most abundant in binding pockets. Significant differences between drug-like and physiological compounds were found. On average, physiological compounds establish with respect to drugs about twice as many hydrogen bonds with protein atoms, whereas drugs rely more on hydrophobic interactions to establish target selectivity. The large number of PDB structures describing homologous proteins in complex with the same ligand made it possible to analyze the conservation of binding pocket residues among homologous protein structures bound to the same ligand, showing that Gly, Glu, Arg, Asp, His, and Thr are more conserved than other amino acids. Also in the cases in which the same ligand is bound to unrelated proteins, the binding pockets showed significant conservation in the residue types. In this case, the probability of co-occurrence of the same amino acid type in the binding pockets could be up to thirteen times higher than that expected on a random basis. The trends identified in this study may provide an useful guideline in the process of drug design and lead optimization.
TL;DR: Functional analysis of M37A mutation in HTLV PR clearly shows that the MET37 specificity and screening of potential inhibitors targeting MET37 is performed by using approved 90% similar HIV PR inhibitor compounds.
Abstract: Retroviruses HTLV-1 and HIV-1 are the primary causative agents of fatal adult T-cell leukemia and acquired immune deficiency syndrome (AIDS) disease. Both retroviruses are similar in characteristics mechanism, and it encodes for protease that mainly involved in the viral replication process. On the basis of the therapeutic success of HIV-1 PR inhibitors, the protease of HTLV-1 is mainly considered as a potential target for chemotherapy. At the same time, structural similarities in both enzymes that originate HIV PR inhibitors can also be an HTLV-1 PR inhibitor. But the expectations failed because of rejection of HIV PR inhibitors from the HTLV-1 PR binding pocket. In this present study, the reason for the HIV PR inhibitor rejection from the HTLV-1 binding site was identified through sequence analysis and molecular dynamics simulation method. Functional analysis of M37A mutation in HTLV PR clearly shows that the MET37 specificity and screening of potential inhibitors targeting MET37 is performed by using approved 90% similar HIV PR inhibitor compounds. From this approach, we report few compounds with a tendency to accept/donate electron specifically to an important site residue MET37 in HTLV-1 PR binding pocket. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: In this article, the authors performed molecular dynamics simulations for the PLP-dependent enzyme tryptophan synthase (TRPS), whose catalytic mechanism features multiple quasi-stable intermediates.
Abstract: The importance of protonation states and proton transfer in pyridoxal 5′-phosphate (PLP)-chemistry can hardly be overstated. Although experimental approaches to investigate pKa values can provide general guidance for assigning proton locations, only static pictures of the chemical species are available. To obtain the overall protein dynamics for the interpretation of detailed enzyme catalysis in this study, guided by information from solid-state NMR, we performed molecular dynamics (MD) simulations for the PLP-dependent enzyme tryptophan synthase (TRPS), whose catalytic mechanism features multiple quasi-stable intermediates. The primary objective of this work is to elucidate how the position of a single proton on the reacting substrate affects local and global protein dynamics during the catalytic cycle. In general, proteins create a chemical environment and an ensemble of conformational motions to recognize different substrates with different protonations. The study of these interactions in TRPS shows that functional groups on the reacting substrate, such as the phosphoryl group, pyridine nitrogen, phenolic oxygen and carboxyl group, of each PLP-bound intermediate play a crucial role in constructing an appropriate molecular interface with TRPS. In particular, the protonation states of the ionizable groups on the PLP cofactor may enhance or weaken the attractions between the enzyme and substrate. In addition, remodulation of the charge distribution for the intermediates may help generate a suitable environment for chemical reactions. The results of our study enhance knowledge of protonation states for several PLP intermediates and help to elucidate their effects on protein dynamics in the function of TRPS and other PLP-dependent enzymes.
TL;DR: Molecular models of Fd docked onto the stromal domain of the cyanobacterial PSI site are constructed here utilizing X‐ray and NMR structures of PSI and Fd, respectively, which predict that these regions interact predominantly with regions of high localized frustration on the PsaC, PsaD, and PsaE chains of PSi.
Abstract: The stromal domain (PsaC, PsaD, and PsaE) of photosystem I (PSI) reduces transiently bound ferredoxin (Fd) or flavodoxin. Experimental structures exist for all of these protein partners individually, but no experimental structure of the PSI/Fd or PSI/flavodoxin complexes is presently available. Molecular models of Fd docked onto the stromal domain of the cyanobacterial PSI site are constructed here utilizing X-ray and NMR structures of PSI and Fd, respectively. Predictions of potential protein-protein interaction regions are based on experimental site-directed mutagenesis and cross-linking studies to guide rigid body docking calculations of Fd into PSI, complemented by energy landscape theory to bring together regions of high energetic frustration on each of the interacting proteins. The results identify two regions of high localized frustration on the surface of Fd that contain negatively charged Asp and Glu residues. This study predicts that these regions interact predominantly with regions of high localized frustration on the PsaC, PsaD, and PsaE chains of PSI, which include several residues predicted by previous experimental studies. Copyright © 2014 John Wiley & Sons, Ltd. Additional supporting information may be found in the online version of this article at the publisher’s web site.
TL;DR: Geometric analyses of subunit coordinates provided a quantitative description of the particle reorganization during maturation, showing that the subunits differentiate from near equivalent environments in the procapsid to strikingly non‐equivalent environments during m maturity.
Abstract: Quasi-equivalent viruses that infect animals and bacteria require a maturation process in which particles transition from initially assembled procapsids to infectious virions. Nudaurelia capensis ω virus (NωV) is a T = 4, eukaryotic, single-stranded ribonucleic acid virus that has proved to be an excellent model system for studying the mechanisms of viral maturation. Structures of NωV procapsids (diameter = 480 A), a maturation intermediate (410 A), and the mature virion (410 A) were determined by electron cryo-microscopy and three-dimensional image reconstruction (cryoEM). The cryoEM density for each particle type was analyzed with a recently developed maximum likelihood variance (MLV) method for characterizing microstates occupied in the ensemble of particles used for the reconstructions. The procapsid and the mature capsid had overall low variance (i.e., uniform particle populations) while the maturation intermediate (that had not undergone post-assembly autocatalytic cleavage) had roughly two to four times the variance of the first two particles. Without maturation cleavage, the particles assume a variety of microstates, as the frustrated subunits cannot reach a minimum energy configuration. Geometric analyses of subunit coordinates provided a quantitative description of the particle reorganization during maturation. Superposition of the four quasi-equivalent subunits in the procapsid had an average root mean square deviation (RMSD) of 3 A while the mature particle had an RMSD of 11 A, showing that the subunits differentiate from near equivalent environments in the procapsid to strikingly non-equivalent environments during maturation. Autocatalytic cleavage is clearly required for the reorganized mature particle to reach the minimum energy state required for stability and infectivity.
TL;DR: CoxyL presented hemagglutinating activity that was specifically inhibited by monosaccharides (D‐glucose, D‐mannose, and α‐methyl‐D‐mannoside) and glycoproteins (ovalbumin and fetuin).
Abstract: Recent studies have shown that lectins are promising tools for use in various biotechnological processes, as well as studies of various pathological mechanisms, isolation, and characterization of glycoconjugates and understanding the mechanisms underlying pathological mechanisms conditions, including the inflammatory response. This study aimed to purify, characterize physicochemically, and predict the biological activity of Canavalia oxyphylla lectin (CoxyL) in vitro and in vivo. CoxyL was purified by a single-step affinity chromatography in Sephadex® G-50 column. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that the pure lectin consists of a major band of 30 kDa (α-chain) and two minor components (β-chain and γ-chain) of 16 and 13 kDa, respectively. These data were further confirmed by electrospray ionization mass spectrometry, suggesting that CoxyL is a typical ConA-like lectin. In comparison with the average molecular mass of α-chain, the partial amino acid sequence obtained corresponds to approximately 45% of the total CoxyL sequence. CoxyL presented hemagglutinating activity that was specifically inhibited by monosaccharides (D-glucose, D-mannose, and α-methyl-D-mannoside) and glycoproteins (ovalbumin and fetuin). Moreover, CoxyL was shown to be thermostable, exhibiting full hemagglutinating activity up to 60°C, and it was pH-sensitive for 1 h, exhibiting maximal activity at pH 7.0. CoxyL caused toxicity to Artemia nauplii and induced paw edema in rats. This biological activity highlights the importance of lectins as important tools to better understand the mechanisms underlying inflammatory responses.
TL;DR: A novel ratiometric fluorescent receptor bearing one phenolic OH and one aldehyde group as recognition sites was synthesized and characterized and showed fast response time, excellent selectivity and reproducibility towards iodide ion detection among the other surveyed anions.
Abstract: A novel ratiometric fluorescent receptor (Z)-2-(4-[diethylamino]-2-hydroxybenzylideneamino) pyridine-3-carbaldehyde (3) bearing one phenolic OH and one aldehyde group as recognition sites was synthesized and characterized. The anion recognition behaviour of receptor 3 was evaluated by various spectroscopic (UV-visible, fluorescence and 1H nuclear magnetic resonance) methods and was validated by computational studies. The receptor showed fast response time, excellent selectivity and reproducibility towards iodide ion detection among the other surveyed anions, with a binding constant of 6.12 × 104 M−1 and a detection limit of 0.24 μM, thus confirming its potential applicability as a fluorescent sensor for iodide Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: It was found that the residues of tryptophan and tyrosine on the ligands have significant contributions for the Fc–ligand binding, while Met252, Ile253, Asn434, His435, and Tyr436 are the key residues of Fc.
Abstract: Immunoglobulin G (IgG) plays an important role in clinical diagnosis and therapeutics. Meanwhile, the consensus binding site (CBS) on the Fc domain of IgG is responsible for ligand recognition, especially for Fc-specific ligands. In this study, molecular simulation methods were used to investigate molecular interactions between the CBS of the Fc domain and seven natural Fc-specific ligands. The analysis on the binding energy of the Fc-ligand complex indicated that hydrophobic interactions provide the main driving force for the Fc-ligand binding processes. The hot spots on the ligands and Fc were identified with the computational alanine scanning approach. It was found that the residues of tryptophan and tyrosine on the ligands have significant contributions for the Fc-ligand binding, while Met252, Ile253, Asn434, His435, and Tyr436 are the key residues of Fc. Moreover, two binding modes based on tryptophan or tyrosine were summarized and constructed according to the pairwise interaction analysis. Guidelines for the rational design of CBS-specific ligands with high affinity and specificity were proposed.
TL;DR: The interaction between azurin (Az) and cytochrome c 551 (CytC551) from Pseudomonas aeruginosa has been studied by surface plasmon resonance and fluorescence quenching as mentioned in this paper.
Abstract: The interaction between azurin (Az) and cytochrome c 551 (CytC551) from Pseudomonas aeruginosa deserves particular interest for both its physiological aspects and their possible applications in bionano devices. Here, the kinetics of the interaction has been studied by surface plasmon resonance and fluorescence quenching. Surface plasmon resonance data have been successfully interpreted by the heterogeneous ligand model, which predicts the existence of two binding sites on the immobilized Az for CytC551 molecules in solution. On the other hand, the fluorescence study indicates the formation of a complex, with the involvement of the lone Az tryptophan (Trp) at position 48. The two different techniques point out the occurrence of an encounter complex between Az and CytC551 that evolves toward the formation of a more stable complex characterized by an equilibrium dissociation constant KD typical of transient interactions.
TL;DR: A knowledge‐driven approach takes advantage of experimentally identified inhibitors against a particular protein target of interest to delineate shape and molecular field properties and use a multilayer perceptron model to predict the biological activity of the test molecules.
Abstract: Prioritization of compounds using inverse docking approach is limited owing to potential drawbacks in its scoring functions. Classically, molecules ranked by best or lowest binding energies and clustering methods have been considered as probable hits. Mining probable hits from an inverse docking approach is very complicated given the closely related protein targets and the chemically similar ligand data set. To overcome this problem, we present here a computational approach using receptor-centric and ligand-centric methods to infer the reliability of the inverse docking approach and to recognize probable hits. This knowledge-driven approach takes advantage of experimentally identified inhibitors against a particular protein target of interest to delineate shape and molecular field properties and use a multilayer perceptron model to predict the biological activity of the test molecules. The approach was validated using flavone derivatives possessing inhibitory activities against principal antimalarial molecular targets of fatty acid biosynthetic pathway, FabG, FabI and FabZ, respectively. We propose that probable hits can be retrieved by comparing the rank list of docking, quantitative-structure activity relationship and multilayer perceptron models. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: The findings suggest that the TPR domain from AtToc64 has higher affinity towards C‐terminal residues of Hsp70 and Hsp90, which are two chaperones known to play import roles in protein import.
Abstract: The study aims to gain insight into the mode of ligand recognition by tetratricopeptide repeat (TPR) domains of chloroplast translocon at the outer envelope of chloroplast (Toc64) and mitochondrial Om64, two paralogous proteins that mediate import of proteins into chloroplast and mitochondria, respectively. Chaperone proteins associate with precursor proteins in the cytosol to maintain them in a translocation competent conformation and are recognized by Toc64 and Om64 that are located on the outer membrane of the target organelle. Heat shock proteins (Hsp70) and Hsp90 are two chaperones, which are known to play import roles in protein import. The C-termini of these chaperones are known to interact with the TPR domain of chloroplast Toc64 and mitochondrial Om64 in Arabidopsis thaliana (At). Using a molecular dynamics approach and binding energy calculations, we identify important residues involved in the interactions. Our findings suggest that the TPR domain from AtToc64 has higher affinity towards C-terminal residues of Hsp70. The interaction occurs as the terminal helices move towards each other enclosing the cradle on interaction of AtHsp70 with the TPR domain. In contrast, the TPR domain from AtOm64 does not discriminate between the C-termini of Hsp70 and Hsp90. These binding affinities are discussed with respect to our knowledge of protein targeting and specificity of protein import into endosymbiotic organelles in plant cells. Copyright © 2014 John Wiley & Sons, Ltd.
TL;DR: Double point mutated WGA (WGA3) exhibits increased specificity towards sialylgalactose, and thus, it can be effectively used in targeted drug delivery and as biological cell marker in cancer therapeutics.
Abstract: Wheat germ agglutinin (WGA) is a plant lectin, which specifically recognizes the sugars NeuNAc and GlcNAc. Mutated WGA with enhanced binding specificity can be used as biomarkers for cancer. In silico mutations are performed at the active site of WGA to enhance the binding specificity towards sialylglycans, and molecular dynamics simulations of 20 ns are carried out for wild type and mutated WGAs (WGA1, WGA2, and WGA3) in complex with sialylgalactose to examine the change in binding specificity. MD simulations reveal the change in binding specificity of wild type and mutated WGAs towards sialylgalactose and bound conformational flexibility of sialylgalactose. The mutated polar amino acid residues Asn114 (S114N), Lys118 (G118K), and Arg118 (G118R) make direct and water mediated hydrogen bonds and hydrophobic interactions with sialylgalactose. An analysis of possible hydrogen bonds, hydrophobic interactions, total pair wise interaction energy between active site residues and sialylgalactose and MM-PBSA free energy calculation reveals the plausible binding modes and the role of water in stabilizing different binding modes. An interesting observation is that the binding specificity of mutated WGAs (cyborg lectin) towards sialylgalactose is found to be higher in double point mutation (WGA3). One of the substituted residues Arg118 plays a crucial role in sugar binding. Based on the interactions and energy calculations, it is concluded that the order of binding specificity of WGAs towards sialylgalactose is WGA3 > WGA1 > WGA2 > WGA. On comparing with the wild type, double point mutated WGA (WGA3) exhibits increased specificity towards sialylgalactose, and thus, it can be effectively used in targeted drug delivery and as biological cell marker in cancer therapeutics. Copyright © 2014 John Wiley & Sons, Ltd.