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Binding Behavior (binding + behavior)
Selected AbstractsEffect of Electronic Structures of Enantiomers of Ruthenium(II) Polypyridyl Complexes on DNA Binding BehaviorsCHINESE JOURNAL OF CHEMISTRY, Issue 8 2010Haimei Luo Abstract A pair of Ru(II) complex enantiomers, , - and , -[Ru(bpy)2(p -mpip)]2+ {bpy=2,2,-bipyridine, p -mpip=2-(4-methylphenyl)imidazo[4,5-f]-1,10-phenanthroline} have been synthesized and structurally characterized. Both experimental results from crystallography, NMR, electrochemistry and theoretical calculations applying the density functional theory (DFT) method based on their crystal structures show that small difference in geometric structure existed can cause a considerable difference in electronic structure between enantiomers. In addition, the binding of the two enantiomers to calf thymus DNA (CT DNA) has been investigated with UV spectroscopy titration and viscosity measurements. It is very rare that the , enantiomer binds to DNA more strongly than the , enantiomer, which can be reasonably explained by their different electronic structures for the first time, suggesting that the dominant factor governing the stereoselectivity of DNA binding of Ru(II) complex may be the different electronic structures of its enantiomers. [source] Determining the Zero-Force Binding Energetics of an Intercalated DNA Complex by a Single-Molecule ApproachCHEMPHYSCHEM, Issue 16 2009Tzu-Sen Yang Prof. Dr. Binding behavior of DNA: A wormlike chain model is applied to determine the zero-force binding energetics of an intercalated DNA complex. A mono-intercalating agent is synthesized which can be very easily inserted into the DNA bases (see figure). This approach is particularly important in nuclear medicine and new radiation therapies. [source] Voltammetric Studies of the Interactions Between Ferrocene-Labeled Glutathione and Proteins in Solution or Immobilized onto SurfaceELECTROANALYSIS, Issue 16 2009Yong Peng Abstract Glutathione (GSH) tagged with a ferrocene (Fc) label at its C-terminal was synthesized via coupling ferrocenyl amine to glutathione using o -(benzotriazol-1-yl)- N,N,N,,N, -tetramethyluronium (HBTU)/1-hydroxybenzotrizole (HOBt). The presence of Fc yielded well defined voltammetric signals, rendering the Fc-tagged GSH (GSH-Fc) suitable for electrochemical studies of GSH binding to other biological species. The interaction of GSH-Fc with bovine serum albumin (BSA) was investigated, and a binding ratio of 1.41±0.06 (GSH-Fc/BSA) and an affinity constant Ka of 6.53±2.01×106,M,1 were determined. These results compare well with those measured by fluorescence using untagged GSH, suggesting that the attachment of Fc to GSH does not significantly perturb the GSH structure and binding behavior. By contrasting the binding behavior to several compounds that are known to conjugate to different domains of BSA, the voltammetric study confirmed that GSH-Fc binds at subdomain IIA of BSA with high affinity. The versatility of GSH-Fc for studying GSH binding to surface-confined proteins was also demonstrated with the GSH binding to electroinactive Zn-metallothionein (Zn7 -MT) through hydrogen binding at the region between the Zn7 -MT , and , domains. [source] Capillary electrophoresis as a probe of enantiospecific interactions between photoactive transition metal complexes and DNAELECTROPHORESIS, Issue 15 2003James P. Schaeper Abstract Recently, we have demonstrated the capacity to separate chiral transition metal (TM) complexes of the type [M(diimine)3]n+ using CE buffers containing chiral tartrate salts. In separate work, several chromium(III)- tris -diimine complexes in particular have been shown to bind enantioselectively with calf-thymus (CT) DNA, and a qualitative assessment of the relative strength and enantiospecificity of this interaction is of significant interest in the characterization of these complexes as potential DNA photocleavage agents. Here, we describe two convenient approaches to investigate such binding behavior using chiral CE. For complexes with lower DNA affinities exhibiting primarily surface binding, DNA itself is used as the chiral resolving agent in the electrophoretic buffer. In this approach, resolution of the TM complexes into their , and , isomers is achieved with the isomer eluting later exhibiting superior binding affinity toward DNA. For more strongly bound TM complexes containing ligands known to intercalate with DNA, the [Cr(diimine)3]3+ complexes are preincubated with oligonucleotide and subsequently enantiomerically resolved in a dibenzoyl- L -tartrate buffer system that facilitates analysis of the unbound TM species only. Differences in isomer binding affinity are distinguished by the relative peak areas of the ,- and ,-isomers, and relative binding strengths of different complexes can be inferred from comparison of the total amount of unbound complex at equivalent DNA/TM ratios. [source] Microfibril-associated glycoprotein-1 binding to tropoelastinFEBS JOURNAL, Issue 14 2004Multiple binding sites, the role of divalent cations Microfibrils and elastin are major constituents of elastic fibers, the assembly of which is dictated by multimolecular interactions. Microfibril-associated glycoprotein-1 (MAGP-1) is a microfibrillar component that interacts with the soluble elastin precursor, tropoelastin. We describe here the adaptation of a solid-phase binding assay that defines the effect of divalent cations on the interactions between MAGP-1 and tropoelastin. Using this assay, a strong calcium-dependent interaction was demonstrated, with a dissociation constant of 2.8 ± 0.3 nm, which fits a single-site binding model. Manganese and magnesium bestowed a weaker association, and copper did not facilitate the protein interactions. Three constructs spanning tropoelastin were used to quantify their relative contributions to calcium-dependent MAGP-1 binding. Binding to a construct spanning a region from the N-terminus to domain 18 followed a single-site binding model with a dissociation constant of 12.0 ± 2.2 nm, which contrasted with the complex binding behavior observed for fragments spanning domains 17,27 and domain 27 to the C-terminus. To further elucidate binding sites around the kallikrein cleavage site of domains 25/26, MAGP-1 was presented with constructs containing C-terminal deletions within the region. Construct M1659, which spans a region from the N-terminus of tropoelastin to domain 26, inclusive, bound MAGP-1 with a dissociation constant of 9.7 ± 2.0 nm, which decreased to 4.9 ± 1.0 nm following the removal of domain 26 (M155n), thus displaying only half the total capacity to bind MAGP-1. These results demonstrate that MAGP-1 is capable of cumulative binding to distinct regions on tropoelastin, with different apparent dissociation constants and different amounts of bound protein. [source] Side chain substitution benchmark for peptide/MHC interactionPROTEIN SCIENCE, Issue 6 2008Bernhard Knapp Abstract The prediction of T-cell epitopes is an essential part in virtual immunology. Apart from sequence-based techniques, which achieve good results but fail to give insight into the binding behavior of a certain peptide binding to a major histocompatibility complex, structure-based approaches are another important technique. An essential step is the correct placement of the side chains for a given peptide in cases where no experimental data for the structure are available. To our knowledge, no benchmark for side chain substitution in the area of HLA has been reported in the literature. Here, we present a comparison of five different tools (SCWRL, SCATD, SPDBV, SCit, IRECS) applicable for side chain substitution. Each tool is tested on 29 different HLA-A2 structures with experimentally known side chain positions. Parts of the benchmark are correctness, reliability, runtime, and usability. For validation, the root mean square deviations between X-ray structures and predicted structures are used. All tools show different strengths and weaknesses. [source] Arm,domain interactions can provide high binding cooperativityPROTEIN SCIENCE, Issue 10 2004Robert Schleif Abstract Peptidyl arms extending from one protein domain to another protein domain mediate many important interactions in biology. A well-studied example of this type of protein,protein interaction occurs between the yeast homeodomain proteins, MAT ,2 and MAT a1, which form a high-affinity heterodimer on DNA. The carboxyl-terminal arm extending from MAT ,2 to MAT a1 has been proposed to produce an allosteric conformational change in the a1 protein that generates a very large increase in the DNA binding affinity of a1. Although early studies lent some support to this model, a more recent crystal structure determination of the free a1 protein argues against any allosteric change. This note presents a thermodynamic argument that accounts for the proteins' binding behavior, so that allosteric conformational changes are not required to explain the large affinity increase. The analysis presented here should be useful in analyzing binding behavior in other systems involving arm interactions. [source] New Inhibitors of the Tat,TAR RNA Interaction Found with a "Fuzzy" Pharmacophore ModelCHEMBIOCHEM, Issue 6 2005Steffen Renner Dipl.-Biol. Abstract TAR RNA is a potential target for AIDS therapy. Ligand-based virtual screening was performed to retrieve novel scaffolds for RNA-binding molecules capable of inhibiting the Tat,TAR interaction, which is essential for HIV replication. We used a "fuzzy" pharmacophore approach (SQUID) and an alignment-free pharmacophore method (CATS3D) to carry out virtual screening of a vendor database of small molecules and to perform "scaffold-hopping". A small subset of 19 candidate molecules were experimentally tested for TAR RNA binding in a fluorescence resonance energy transfer (FRET) assay. Both methods retrieved molecules that exhibited activities comparable to those of the reference molecules acetylpromazine and chlorpromazine, with the best molecule showing ten times better binding behavior (IC50=46 ,M). The hits had molecular scaffolds different from those of the reference molecules. [source] ADLOC: An Aptamer-Displacement Assay Based on Luminescent Oxygen ChannelingCHEMISTRY - A EUROPEAN JOURNAL, Issue 36 2010Dipl.-Chem. Abstract Functional nucleic acids, such as aptamers and allosteric ribozymes, can sense their ligands specifically, thereby undergoing structural alterations that can be converted into a detectable signal. The direct coupling of molecular recognition to signal generation enables the production of versatile reporters that can be applied as molecular probes for various purposes, including high-throughput screening. Here we describe an unprecedented type of a nucleic acid-based sensor system and show that it is amenable to high-throughput screening (HTS) applications. The approach detects the displacement of an aptamer from its bound protein partner by means of luminescent oxygen channeling. In a proof-of-principle study we demonstrate that the format is feasible for efficient identification of small drug-like molecules that bind to a protein target, in this case to the Sec7 domain of cytohesin. We extended the approach to a new cytohesin-specific single chain DNA aptamer, C10.41, which exhibits a similar binding behavior to cytohesins but has the advantage of being more stable and easier to synthesize and to modify than the RNA-aptamer M69. The results obtained with both aptamers indicate the general suitability of the aptamer-displacement assay based on luminescent oxygen channelling (ADLOC) for HTS. We also analyzed the potential for false positive hits and identified from a library of 18,000 drug-like small molecules two compounds as strong singlet-oxygen quenchers. With full automation and the use of commercially available plate readers, we estimate that the ADLOC-based assay described here could be used to screen at least 100,000 compounds per day. [source] Tuning Specific Biomolecular Interactions Using Electro-Switchable Oligopeptide SurfacesADVANCED FUNCTIONAL MATERIALS, Issue 16 2010Chun L. Yeung Abstract The ability to regulate biomolecular interactions on surfaces driven by an external stimuli is of great theoretical interest and practical impact in the biomedical and biotechnology fields. Herein, a new class of responsive surfaces that rely on electro-switchable peptides to control biomolecular interactions on gold surfaces is presented. This system is based upon the conformational switching of positively charged oligolysine peptides that are tethered to a gold surface, such that bioactive molecular moieties (biotin) incorporated on the oligolysines can be reversibly exposed (bio-active state) or concealed (bio-inactive state) on demand, as a function of surface potential. The dynamics of switching the biological properties is studied by observing the binding events between biotin and fluorescently labeled NeutrAvidin. Fluorescence microscope images and surface plasmon resonance spectral data clearly reveal opposite binding behaviors when +0.3 V or ,0.4 V vs. SCE are applied to the surface. High fluorescence intensities are observed for an applied positive potential, while minimal fluorescence is detected for an applied negative potential. Surface plasmon resonance spectroscopy (SPR) results provided further evidence that NeutrAvidin binding to the surface is controlled by the applied potential. A large SPR response is observed when a positive potential is applied on the surface, while a negative applied potential induces over 90% reduction in NeutrAvidin binding. [source] Cooperative and Competitive Binding in Synergistic Mixtures of Thermobifida fuscaCellulases Cel5A, Cel6B, and Cel9ABIOTECHNOLOGY PROGRESS, Issue 4 2002Tina Jeoh Synergism between cellulases facilitates efficient hydrolysis of microcrystalline cellulose. We hypothesize that the effects of synergism, observed as enhanced extents of hydrolysis, are related to cellulase binding to the substrate in mixtures. In this study, direct measurements of bound concentrations of fluorescence-labeled T. fuscaCel5A, Cel6B, and Cel9A on bacterial microcrystalline cellulose were used to study binding behaviors of cellulases in binary component reactions. The accuracy of the determination of fluorescence-labeled cellulase concentrations in binary component mixtures was in the range of 7,9%. Data at 5 °C show that binding levels of cellulases in mixture reactions are only 22,70% of the binding levels in single component reactions. At 50 °C, however, most of the cellulase components in the same mixtures bound to extents of 40,126% higher than in the corresponding single component reactions. The degrees of synergistic effect (DSE) observed for the reactions at 50 °C were greater than 1, indicating that the components in the mixture acted synergistically, whereas DSE < 1 was generally observed for the reactions at 5 °C indicating anti-synergistic behavior. Degrees of synergistic binding (DSB) were also calculated, where anti-synergistic mixtures had DSB < 1 and synergistic mixtures had DSB>1. We conclude that the lower extents of binding at 5 °C are due to competition for binding sites by the cellulase components in the mixtures and the enhanced binding extents at 50 °C are due to increased availability of binding sites on the substrates brought about by the higher extents of hydrolysis. [source] | |||||||||||||