Significant Role In Binding Research Articles

Unveiling crucial amino acids in the carbohydrate recognition domain of a viral protein through a structural bioinformatic approach.

Carbohydrate binding modules (CBMs) are protein domains that typically reside near catalytic domains, increasing substrate-protein proximity by constraining the conformational space of carbohydrates. Due to the flexibility and variability of glycans, the molecular details of how these protein regions recognize their target molecules are not always fully understood. Computational methods, including molecular docking and molecular dynamics simulations, have been employed to investigate lectin-carbohydrate interactions. In this study, we introduce a novel approach that integrates multiple computational techniques to identify the critical amino acids involved in the interaction between a CBM located at the tip of bacteriophage J-1's tail and its carbohydrate counterparts. Our results highlight three amino acids that play a significant role in binding, a finding we confirmed through in vitro experiments. By presenting this approach, we offer an intriguing alternative for pinpointing amino acids that contribute to protein-sugar interactions, leading to a more thorough comprehension of the molecular determinants of protein-carbohydrate interactions.

Non-canonical proline-tyrosine interactions with multiple host proteins regulate Ebola virus infection.

The Ebola virus VP30 protein interacts with the viral nucleoprotein and with host protein RBBP6 via PPxPxY motifs that adopt non-canonical orientations, as compared to other proline-rich motifs. An affinity tag-purification mass spectrometry approach identified additional PPxPxY-containing host proteins hnRNP L, hnRNPUL1, and PEG10, as VP30 interactors. hnRNP L and PEG10, like RBBP6, inhibit viral RNA synthesis and EBOV infection, whereas hnRNPUL1 enhances. RBBP6 and hnRNP L modulate VP30 phosphorylation, increase viral transcription, andexert additive effects on viral RNA synthesis. PEG10 has more modest inhibitory effects on EBOV replication. hnRNPUL1 positively affects viral RNA synthesis but in a VP30-independent manner. Binding studies demonstrate variable capacity of the PPxPxY motifs from these proteins to bind VP30, define PxPPPPxY as an optimal binding motif, and identify the fifth proline and the tyrosine as most critical for interaction. Competition binding and hydrogen-deuterium exchange mass spectrometry studies demonstrate that each protein binds a similar interface on VP30. VP30 therefore presents a novel proline recognition domain that is targeted by multiple host proteins to modulate viral transcription.

Thermodynamic and mechanistic analytical effect of albumin coated gold nanosystems for antibiotic drugs binding and interaction with deoxyribonucleic acid

Gold nanoclusters offer useful photophysical properties and are extensively used as fluorescent labels in biomedical applications. Conventional drug administrative methods can face disadvantages such as poor solubility and low bioavailability of drugs. In order to overcome these limitations, the need of drug delivery vehicles arises. Here we report quantitative insights into the interaction of antibiotic drugs tetracycline, oxytetracycline, and rolitetracycline with protein (HSA) coated gold nanoclusters (PCGNCs) and that of (PCGNCs + drug) complex with deoxyribonucleic acid to explore drug transport abilities and potential in drug delivery. A combined calorimetric and spectroscopic approach has been employed to meet objectives. Exothermic heat of interaction suggest that polar association takes place between tetracycline or oxytetracycline and PCGNCs whereas rolitetracycline with pyrrolidinomethyl group shows endothermic heat of interaction. PCGNCs offer improved binding to drugs as compared to that with just the protein. These drug bound PCGNCs are further observed to undergo strong association with DNA in intercalation mode. Exothermic heats of interaction of (PCGNCs + tetracycline) and (PCGNCs + oxytetracycline) with DNA reveal that polar interactions play a significant role in binding whereas (PCGNCs + rolitetracycline) favors entropy driven process. Thermodynamic information associated with structural properties of the drugs and their carriers provide essential guidelines for developing effective drug delivery systems.

Comparative Docking Studies: A Drug Design Tool for Some Pyrazine- Thiazolidinone Based Derivatives for Anti-HIV Activity.

Acquired immunodeficiency Syndrome (AIDS) is caused by Human immunodeficiency virus type 1 (HIV-1). Pyrazine and Thiazolidinone pharmacophore has diverse biological activities including anti HIV activity. To study binding behavior of Pyrazine- thiazolidinone derivatives on four different crystal structures of HIV- 1RT.These molecules which were already reported as anti-TB were investigated for dual activity as Anti-HIV and Anti-TB. In the present study we describe a comparative docking study of twentythree derivatives of N-(4-oxo-2 substituted thiazolidin-3-yl) pyrazine-2-carbohydrazide. Binding pattern of these derivatives was gauged by molecular docking studies on four different receptors bearing PDB code 1ZD1, 1RT2, 1FKP and 1FK9 of HIV-RT enzyme using V. Life MDS software Genetic algorithm docking method. The studies revealed hydrogen bonds, hydrophobic interaction and pi-pi interactions playing significant role in binding of the molecules to the enzyme. Most of the molecules have shown good dock score and binding energy with anti-HIV receptors but Molecules 13 and 14 have potential to act as anti-tubercular and Anti HIV and hence can be further explored for dual activity.

Comparative Study of Various Non-Nucleoside Reverse Transcriptase Inhibitors on Different Reverse Transcriptase Enzyme

Abstract: Context: Acquired immunodeficiency Syndrome (AIDS) is caused by Human immunodeficiency virus type 1 (HIV-1). 4-Thiazolidone nulecus is the target pharmacophore which have diverse biological activities including anti HIV activity. Aim: To study binding behavior of thiazolidinone derivatives on four different crystal structures of HIV- 1RT. Material and Method: Binding pattern of some thiazolidinone derivatives was gauged by molecular docking studies on four different receptors bearing PDB code 1ZD1, 1RT2, 1KLM, 1FKP of HIV–RT enzyme using V. Life MDS software. Result and Discussion: The studies revealed hydrogen bonds, hydrophobic interaction and pi-pi interactions playing significant role in binding of the molecules to the enzyme. Conclusion: Interactions, binding energy and dock score of molecule 6 was comparable with the standard drugs. Key words: Molecular Docking, Thiazolidinone, HIV, NNRTI.

A Survey of DDX21 Activity During Rev/RRE Complex Formation

HIV-1 requires a specialized nuclear export pathway to transport unspliced and partially spliced viral transcripts to the cytoplasm. Central to this pathway is the viral protein Rev, which binds to the Rev response element in stem IIB located on unspliced viral transcripts and subsequently oligomerizes in a cooperative manner. Previous work identified a number of cellular DEAD-box helicases as in vivo binding partners of Rev, and siRNA experiments indicated a functional role for many in the HIV replication cycle. Two DEAD-box proteins, DDX1 and DDX3, had previously been shown to play a role in HIV pathogenesis. In this study, another protein identified in that screen, DDX21, is tested for protein and RNA binding and subsequent enzymatic activities in the context of the Rev/RRE pathway. We found that DDX21 can bind to the RRE with high affinity, and this binding stimulates ATPase activity with an enzymatic efficiency similar to DDX1. Furthermore, DDX21 is both an ATP-dependent and ATP-independent helicase, and both ATPase and ATP-dependent helicase activities are inhibited by Rev in a dose-dependent manner, although ATP-independent helicase activity is not. A conserved binding interaction between DDX protein's DEAD domain and Rev was identified, with Rev's nuclear diffusion inhibitory signal motif playing a significant role in binding. Finally, DDX21 was shown to enhance Rev binding to the RRE in a manner similar to that previously described for DDX1, although DDX3 does not. These data indicate that DDX1 and DDX21 have similar biochemical activities with regard to the Rev/RRE system, while DDX3 differs.

Fractionation of Selected Heavy Metals in Agricultural Soils / Frakcjonowanie Wybranych Metali Ciężkich W Glebach Uprawnych

Abstract The content of trace elements in soils varies widely and their mobility and availability depends not only on the total content but also on the form of in which these elements occur. The aim of this study was to determine the total content of nickel, lead, zinc and copper in soils used for agriculture, and assess the mobility and phytoavailability of these metals against a background of physical and chemical properties of these soils. In samples taken from three soil profiles (Phaeozem and 2 Fluvisols) the contents of Ni, Pb, Zn and Cu were determined using atomic absorption spectroscopy in the solutions obtained according to the protocol of modified BCR sequential extraction procedure supplemented with aqua regia digestion. The total content of the analyzed metals in most cases corresponded to the natural values, often not exceeding the geochemical background level. It was only in the one profile of the Fluvisols (Endogleyic Fluvisol) that a higher concentration of zinc and lead was noticed (especially in the surface horizon), slightly exceeding the legal limit. Among the studied metals the lowest phytoavailability was characterized by copper (exchangeable forms on average 4.73% of the total), and the highest by zinc (11.49%). Nickel was the most permanently bound with soil solid phase, and its content in the residual fraction reached 84.46% of the total. Approximately a half of the total lead content was determined as a fraction bound with iron and manganese oxides, while in the case of this metal a significant role in binding of this metal was playing organic matter (fraction bound with organic matter and sulphides - an average of 27.5%). Significant role in the binding of all investigated metals was credited to iron and manganese compounds.

BINDING AND SEGMENTATION VIA A NEURAL MASS MODEL TRAINED WITH HEBBIAN AND ANTI-HEBBIAN MECHANISMS

Synchronization of neural activity in the gamma band, modulated by a slower theta rhythm, is assumed to play a significant role in binding and segmentation of multiple objects. In the present work, a recent neural mass model of a single cortical column is used to analyze the synaptic mechanisms which can warrant synchronization and desynchronization of cortical columns, during an autoassociation memory task. The model considers two distinct layers communicating via feedforward connections. The first layer receives the external input and works as an autoassociative network in the theta band, to recover a previously memorized object from incomplete information. The second realizes segmentation of different objects in the gamma band. To this end, units within both layers are connected with synapses trained on the basis of previous experience to store objects. The main model assumptions are: (i) recovery of incomplete objects is realized by excitatory synapses from pyramidal to pyramidal neurons in the same object; (ii) binding in the gamma range is realized by excitatory synapses from pyramidal neurons to fast inhibitory interneurons in the same object. These synapses (both at points i and ii) have a few ms dynamics and are trained with a Hebbian mechanism. (iii) Segmentation is realized with faster AMPA synapses, with rise times smaller than 1 ms, trained with an anti-Hebbian mechanism. Results show that the model, with the previous assumptions, can correctly reconstruct and segment three simultaneous objects, starting from incomplete knowledge. Segmentation of more objects is possible but requires an increased ratio between the theta and gamma periods.

Binding of novel fullerene inhibitors to HIV-1 protease: insight through molecular dynamics and molecular mechanics Poisson–Boltzmann surface area calculations

The objectives of this study include the design of a series of novel fullerene-based inhibitors for HIV-1 protease (HIV-1 PR), by employing two strategies that can also be applied to the design of inhibitors for any other target. Additionally, the interactions which contribute to the observed exceptionally high binding free energies were analyzed. In particular, we investigated: (1) hydrogen bonding (H-bond) interactions between specific fullerene derivatives and the protease, (2) the regions of HIV-1 PR that play a significant role in binding, (3) protease changes upon binding and (4) various contributions to the binding free energy, in order to identify the most significant of them. This study has been performed by employing a docking technique, two 3D-QSAR models, molecular dynamics (MD) simulations and the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method. Our computed binding free energies are in satisfactory agreement with the experimental results. The suitability of specific fullerene derivatives as drug candidates was further enhanced, after ADMET (absorption, distribution, metabolism, excretion and toxicity) properties have been estimated to be promising. The outcomes of this study revealed important protein-ligand interaction patterns that may lead towards the development of novel, potent HIV-1 PR inhibitors.

Cell wall component and mycotoxin moieties involved in the binding of fumonisin B1and B2by lactic acid bacteria

The ability of lactic acid bacteria (LAB) to bind fumonisins B1 and B2 (FB1, FB2) in fermented foods and feeds and in the gastrointestinal tract could contribute to decrease their bioavailability and toxic effects on farm animals and humans. The aim of this work was to identify the bacterial cell wall component(s) and the functional group(s) of FB involved in the LAB-FB interaction. The effect of physicochemical, enzymatic and genetic treatments of bacteria and the removal/inactivation of the functional groups of FB on toxin binding were evaluated. Treatments affecting the bacterial wall polysaccharides, lipids and proteins increased binding, while those degrading peptidoglycan (PG) partially decreased it. In addition, purified PG from Gram-positive bacteria bound FB in a manner analogue to that of intact LAB. For FB, tricarballylic acid (TCA) chains play a significant role in binding as hydrolysed FB had less affinity for LAB. Peptidoglycan and TCA are important components of LAB and FB, respectively, involved in the binding interaction. Lactic acid bacteria binding efficiency seems related to the peptide moiety structure of the PG. This information can be used to select probiotics with increased FB binding efficiency.

Differential contributions of recognition factors of two plant lectins – Amaranthus caudatus lectin and Arachis hypogea agglutinin, reacting with Thomsen-Friedenreich disaccharide (Galβ1–3GalNAcα1–Ser/Thr)

Previous reports on the carbohydrate specificities of Amaranthus caudatus lectin (ACL) and peanut agglutinin (PNA, Arachis hypogea) indicated that they share the same specificity for the Thomsen-Friedenreich ( T α, Galβ1–3GalNAcα1–Ser/Thr) glycotope, but differ in monosaccharide binding – GalNAc ≫ Gal (inactive) for ACL; Gal ≫ GalNAc (weak) with respect to PNA. However, knowledge of the recognition factors of these lectins was based on studies with a small number monosaccharides and T-related oligosaccharides. In this study, a wider range of interacting factors of ACL and PNA toward known mammalian structural units, natural polyvalent glycotopes and glycans were examined by enzyme-linked lectinosorbent and inhibition assays. The results indicate that the main recognition factors of ACL, GalNAc was the only monosaccharide recognized by ACL as such, its polyvalent forms (poly GalNAcα1–Ser/Thr, Tn in asialo OSM) were not recognized much better. Human blood group precursor disaccharides Galβ1–3/4GlcNAcβ ( I β/ II β) were weak ligands, while their clusters (multiantennary II β) and polyvalent forms were active. The major recognition factors of PNA were a combination of α or β anomers of T disaccharide and their polyvalent complexes. Although I β/ II β were weak haptens, their polyvalent forms played a significant role in binding. From the 50% molar inhibition profile, the shape of the ACL combining site appears to be a cavity type and most complementary to a disaccharide of Galβ1–3GalNAc ( T), while the PNA binding domain is proposed to be Galβ1–3GalNAcα or β1- as the major combining site with an adjoining subsite (partial cavity type) for a disaccharide, and most complementary to the linear tetrasaccharide, Galβ1–3GalNAcβ1–4Galβ1–4Glc ( T β1-4 L, asialo GM 1 sequence). These results should help us understand the differential contributions of polyvalent ligands, glycotopes and subtopes for the interaction with these lectins to binding, and make them useful tools to study glycosciences, glycomarkers and their biological functions.

Degradation of trinitrotoluene in contaminated soils as affected by its initial concentrations and its binding to soil organic matter fractions

Trinitrotoluene (TNT), a nitroaromatics, is a major pollutant in explosive contaminated soils. Present study reports the effect of initial concentration of TNT on its degradation kinetics in soils. Soils from two contaminated sites viz. Clausthal and Elsnig, Germany, were mixed with an uncontaminated reference soil to get different initial concentrations (mg/kg) viz Clausthal-1 (54.29), Clausthal-2 (30.86), Clausthal-3 (7.05) Elsnig-1 (879.67), Elsnig-2 (86.43); Elsnig-3 (8.16) and Elsnig-4 (0.99) and were spiked with ring UL-14C-TNT (100KBq/50g soil). Except Elsnig-1 and Elsnig-2 soils, TNT degraded at same rate in all the soils. Higher persistence of TNT in Elsnig-1 and Elsnig-2 soils appears to be due to higher initial concentrations of nitroaromatics which may be toxic to soil microorganisms. 2-Amino-4,6-dinitrotoluene (2-ADNT) and 4-amino-2,6-dinitrotoluene (4-ADNT) were recovered as major metabolites of TNT. Distribution of bound 14C-activity in different soil organic matter (SOM) fractions (humic acid, fulvic acid and humin) was assayed by alkali extraction of solvent extracted Clausthal-1 and Elsnig-1 soils. Results indicate that humic acid accounted for maximum bound activity followed by fulvic acid and humin fractions. Expressing 14C-activity bound/unit weight of organic carbon content of SOM fractions showed that 3 times more 14C-activity was bound to Elsnig humic acid than Clausthal humic acid. Similarly, activity associated with Elsnig fulvic acid was 7 times higher than that of Clausthal fulvic acid suggesting that chemical nature of SOM fractions plays a significant role in binding of contaminants.

Exploring the sequence space of a DNA aptamer using microarrays

The relationship between sequence and binding properties of an aptamer for immunoglobulin E (IgE) was investigated using custom DNA microarrays. Single, double and some triple mutations of the aptamer sequence were created to evaluate the importance of specific base composition on aptamer binding. The majority of the positions in the aptamer sequence were found to be immutable, with changes at these positions resulting in more than a 100-fold decrease in binding affinity. Improvements in binding were observed by altering the stem region of the aptamer, suggesting that it plays a significant role in binding. Results obtained for the various mutations were used to estimate the information content and the probability of finding a functional aptamer sequence by selection from a random library. For the IgE-binding aptamer, this probability is on the order of 10−10 to 10−9. Results obtained for the double and triple mutations also show that there are no compensatory mutations within the space defined by those mutations. Apparently, at least for this particular aptamer, the functional sequence space can be represented as a rugged landscape with sharp peaks defined by highly constrained base compositions. This makes the rational optimization of aptamer sequences using step-wise mutagenesis approaches very challenging.

Binding mode and affinity studies of DNA-binding agents using topoisomerase I DNA unwinding assay

A topoisomerase I DNA unwinding assay has been used to determine the relative DNA-binding affinities of a model pair of homologous naphthalene diimides. Binding affinity data were corroborated using calorimetric (ITC) and spectrophotometric (titration and T m) studies, with substituent size playing a significant role in binding. The assay was also used to investigate the mode of binding adopted by several known DNA-binding agents, including SYBR Green and PicoGreen. Some of the compounds exhibited unexpected binding modes.

Sweet talking–cellular carbohydrates and epithelial repair

Sweet talking–cellular carbohydrates and epithelial repair

Copper complexes as chemical nucleases

Redox active mononuclear and binuclear copper(II) complexes have been prepared and structurally characterized. The complexes have planar N-donor heterocyclic bases like 1,10-phenanthroline (phen), dipyridoquinoxaline (dpq) and dipyridophenazine (dppz) ligands that are suitable for intercalation to B-DNA. Complexes studied for nuclease activity have the formulations [Cu(dpq)2(H2O)] (ClO4)2.H2O (1), [CuL(H2O)2(μ-ox)](ClO4)2 (L = bpy,2; phen,3; dpq,4; and dppz,5) and [Cu(L)(salgly)] (L = bpy,6; phen,7; dpq,8; and dppz,9), where salgly is a tridentate Schiff base obtained from the condensation of glycine and salicylaldehyde. The dpq complexes are efficient DNA binding and cleavage active species. The dppz complexes show good binding ability but poor nuclease activity. The cleavage activity of thebis-dpq complex is significantly higher than thebis-phen complex of copper(II). The nuclease activity is found to be dependent on the intercalating nature of the complex and on the redox potential of the copper(II)/copper(I) couple. The ancillary ligand plays a significant role in binding and cleavage activity.

Mutational analysis of metallo-beta-lactamase CcrA from Bacteroides fragilis.

In an effort to evaluate the roles of Lys184, Asn193, and Asp103 in the binding and catalysis of metallo-beta-lactamase CcrA from Bacteroides fragilis, site-directed mutants of CcrA were generated and characterized using metal analyses, CD spectroscopy, and kinetic studies. Three Lys184 mutants were generated where the lysine was replaced with alanine, leucine, and glutamate, and the analysis of these mutants indicates that Lys184 is not greatly involved in binding of cephalosporins to CcrA; however, this residue does have a significant role in binding of penicillin G. Three Asn193 mutants were generated where the asparagine was replaced with alanine, leucine, and aspartate, and these mutants exhibited <4-fold decrease in k(cat), suggesting that Asn193 does not play a large role in catalysis. However, stopped-flow visible kinetic studies showed that the Asn193 mutants exhibit a slower substrate decay rate and no change in the product formation rate as compared with wild-type CcrA. These results support the proposed role of Asn193 in interacting with and activating substrate during catalysis. Two Asp103 mutants were generated where the aspartate was replaced with serine and cysteine. The D103C and D103S mutants bind the same amount of Zn(II) as wild-type CcrA and exhibited a 10(2)-fold and 10(5)-fold decrease in activity, respectively. Results from solvent isotope, proton inventory, and rapid-scanning visible studies suggest that Asp103 plays a role in generating the enzyme intermediate but does not donate a proton to the enzyme intermediate during the rate-limiting step of the catalytic mechanism.

Thermodynamic evaluation of binding interactions in the methionine repressor system of Escherichia coli using isothermal titration calorimetry.

The binding interactions of the methionine repressor protein, MetJ, from Escherichia coli with its cognate, metbox DNA sequence and corepressor S-adenosylmethionine were examined using calorimetric methods. A detailed thermodynamic characterization of this system which exhibits the recently reported (beta alpha alpha)2 binding motif provides values for delta G, delta H, and delta S for each step in the repressor binding cycle. These studies show that, in the presence of corepressor, MetJ binds to a single metbox operator site with delta G = -7.7 kcal.mol-1, whereas in the absence of corepressor, the free energy of interaction with a single site is -5.8 kcal.mol-1. Cooperative interactions between two repressor molecules bound to two adjacent sites contribute an additional free energy of -1.3 kcal.mol-1 to binding at the second site. Binding is enthalpically unfavorable in the absence of the corepressor with delta H = +2.6 kcal.mol-1 but becomes exothermic with delta H = -4.6 kcal.mol-1 when corepressor is present. The heat capacity for the system decreases significantly by delta Cp = -290 cal.mol-1.K-1 on a per site basis when the protein binds to DNA, and interactions between repressor molecules bound to adjacent sites contribute a delta Cp = -800 cal.mol-1.K-1, indicating that solvent exclusion plays a significant role in binding in this system. The corepressor binds to the unbound repressor protein with a free energy of delta G = -6.0 kcal.mol-1 and to the MetJ-operator complex with delta G = -6.95 kcal.mol-1. Repressor binding to random-sequence DNA was estimated to occur with a free energy of -5.7 kcal.mol-1 in the presence of corepressor. These data clearly indicate that MetJ repressor dimer binds specifically to the central region of its 8 bp cognate metbox operator but recognizes partial operator sequences as short as 6 bp. Cooperativity in binding of adjacent MetJ dimers to a double metbox sequence is demonstrated to be important in determining the energetics of the interaction. Finally, the corepressor S-adenosylmethionine enhances the affinity of MetJ for its recognition site DNA by a factor of 25 and contributes significantly to the net exothermicity of repressor binding.

Ligand binding of bovine carboxypeptidase B. IV. Oligopeptide substrates and extended active center.

Catalytic and binding properties of bovine carboxypeptidase B were studied by kinetic and affinity chromatographic methods both using several oligopeptides as substrates or immobilized ligands. These oligopeptides contained either arginines or phenylalanines at carboxy termini as well as phenylalanyl residues in one of the other positions. The chromatographic studies showed that the phenylalanyl residues in endo-positions play a significant role in binding of the immobilized peptides to the enzyme, while the kinetics studies indicated further that the presence of an internal hydrophobic residue in a substrate was advantageous for the catalytic release of the carboxyl terminal residue from the substrate. These observations support the supposition that the enzyme has an extended active center which contains an extended hydrophobic binding site. Several hydrophobic compounds, which have been shown to act as activators in dipeptide substrate hydrolyses, showed inhibitory effect on hydrolyses of oligopeptide substrates. This observation suggests that these hydrophobic compounds bind to a portion of the hydrophobic site in the active center.