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Ligand Interactions (ligand + interaction)
Selected AbstractsDrug-induced apoptosis in osteosarcoma cell lines is mediated by caspase activation independent of CD95-receptor/ligand interactionJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 1 2000J. Fellenberg Osteosarcoma is one of the most common primary malignant tumors of bone. Treatment of this tumor with systemic chemotherapy dramatically improves the prognosis, although the molecular mechanisms involved in the drug action are poorly understood. In chemosensitive leukaemic T cells and certain solid tumors. cytotoxic drugs mediate the induction of apoptosis by activation of the CD95/APO-1/Fas system. Triggering of the corresponding signaling pathway may involve CD95-receptor/ligand interaction, activation of caspases, or alterations in mitochondrial function. The purpose of our study was to determine if similar mechanisms are involved in the chemosensitivity of osteosarcomas. We found that cytotoxic drugs induce characteristic biochemical and morphological alterations related to apoptosis in osteosarcoma cell lines, including activation of caspases and disturbance of mitochondrial function. However, drug treatment did not result in activation of CD95-receptor or CD95-ligand mRNA. In addition, drug-induced apoptosis was blocked by caspase inhibitors but not by inhibition of CD95-ligand action, indicating a CD95-receptor/ligand-independent mechanism in osteosarcoma cell lines. [source] Book Review: Protein,Ligand Interactions,From Molecular Recognition to Drug Design Methods.CHEMBIOCHEM, Issue 11 2003Edited by Hans-Joachim Böhm, Gisbert Schneider No abstract is available for this article. [source] Multivalent Drug Design and Inhibition of Cholera Toxin by Specific and Transient Protein,Ligand InteractionsCHEMICAL BIOLOGY & DRUG DESIGN, Issue 5 2008Jiyun Liu Multivalent inhibitors of the cholera toxin B pentamer are potential therapeutic drugs for treating cholera and serve as models for demonstrating multivalent ligand effects through a structure-based approach. A crucial yet often overlooked aspect of multivalent drug design is the length, rigidity and chemical composition of the linker used to connect multiple binding moieties. To specifically study the role of chemical linkers in multivalent ligand design, we have synthesized a series of compounds with one and two binding motifs connected by several different linkers. These compounds have affinity for and potency against the cholera toxin B pentamer despite the fact that none can simultaneously bind two toxin receptor sites. Results from saturation transfer difference NMR reveal transient, non-specific interactions between the cholera toxin and linker groups contribute significantly to overall binding affinity of monovalent compounds. However, the same random protein,ligand interactions do not appear to affect binding of bivalent molecules. Moreover, the binding affinities and potencies of these ,non-spanning' bivalent ligands appear to be wholly independent of linker length. Our detailed analysis identifies multiple effects that account for the improved inhibitory potencies of bivalent ligands and suggest approaches to further improve the activity of this class of compounds. [source] Photo-Cross-Linked Small-Molecule Microarrays as Chemical Genomic Tools for Dissecting Protein,Ligand InteractionsCHEMISTRY - AN ASIAN JOURNAL, Issue 6 2006Naoki Kanoh Dr. Abstract We have developed a unique photo-cross-linking approach for immobilizing a variety of small molecules in a functional-group-independent manner. Our approach depends on the reactivity of the carbene species generated from trifluoromethylaryldiazirine upon UV irradiation. It was demonstrated in model experiments that the photogenerated carbenes were able to react with every small molecule tested, and they produced multiple conjugates in most cases. It was also found in on-array immobilization experiments that various small molecules were immobilized, and the immobilized small molecules retained their ability to interact with their binding proteins. With this approach, photo-cross-linked microarrays of about 2000 natural products and drugs were constructed. This photo-cross-linked microarray format was found to be useful not merely for ligand screening but also to study the structure,activity relationship, that is, the relationship between the structural motif (or pharmacophore) found in small molecules and its binding affinity toward a protein, by taking advantage of the nonselective nature of the photo-cross-linking process. [source] Investigation of Protein,Ligand Interactions by Mass SpectrometryCHEMMEDCHEM, Issue 4 2007Andrea Sinz Prof. Abstract The rate of drug discovery is greatly dependent on the development and improvement of rapid and reliable analytical methods that allow screening for protein,ligand interactions. The solution-based methods for investigating protein,ligand interactions by mass spectrometry (MS), which are discussed in this paper, are hydrogen/deuterium exchange of protein backbone amide hydrogens, and photoaffinity labeling. Moreover, MS analysis of intact noncovalent protein,ligand complexes is described. Fourier transform ion cyclotron resonance mass spectrometry (FTICR,MS) with its ultra-high resolution and excellent mass accuracy is also considered herein as it is gaining increasing popularity for a mass spectrometric investigation of protein,ligand interactions. [source] Two-Dimensional Self-Assembly of a Porphyrin,Polypyridyl Ruthenium(II) Hybrid on HOPG Surface through Metal,Ligand InteractionsCHEMPHYSCHEM, Issue 9 2010Aimei Gao Dr. Abstract The synthesis and self-assembly behavior of porphyrin,polypyridyl ruthenium(II) hybrid, which consists of a flexible alkyl chain attached with two conjugated moieties is described. The electronic absorption spectrum and emission spectra show that the [C8 -TPP-(ip)Ru(phen)2](ClO4)2, abbreviated as (C8ip)TPPC has optical properties. Scanning tunneling microscopy (STM) studies found that the ,,, interaction and metal,ligand interaction allow (C8ip)TPPC to form self-assembled structure and have an edge-on orientation on the highly oriented pyrolytic graphite (HOPG) surface. The multidentate structure in (C8ip)TPPC molecules act as linkers between the molecules and form metal,ligand coordination, which forces the assembly process in the direction of stable columnar arrays. In addition, although the sample was stored for two months in ambient conditions, STM experiments showed that the order of (C8ip)TPPC self-assembly only slightly decreased which indicates that the self-assembled monolayer is stable. This work demonstrates that introducing a metal-ligand in the porphyrin-polypyridyl compound is a useful strategy to obtain novel surface assemblies. [source] A Novel Methodological Approach for the Analysis of Host,Ligand Interactions,CHEMPHYSCHEM, Issue 2 2007Daniela Strat Dr. Abstract Traditional analysis of drug-binding data relies upon the Scatchard formalism. These methods rely upon the fitting of a linear equation providing intercept and gradient data that relate to physical properties, such as the binding constant, cooperativity coefficients and number of binding sites. However, the existence of different binding modes with different binding constants makes the implementation of these models difficult. This article describes a novel approach to the binding model of host,ligand interactions by using a derived analytical function describing the observed signal. The benefit of this method is that physically significant parameters, that is, binding constants and number of binding sites, are automatically derived by the use of a minimisation routine. This methodology was utilised to analyse the interactions between a novel antitumour agent and DNA. An optical spectroscopy study confirms that the pentacyclic acridine derivative (DH208) binds to nucleic acids. Two binding modes can be identified: a stronger one that involves intercalation and a weaker one that involves oriented outer-sphere binding. In both cases the plane of the bound acridine ring is parallel to the nucleic acid bases, orthogonal to the phosphate backbone. Ultraviolet (UV) and circular dichroism (CD) data were fitted using the proposed model. The binding constants and the number of binding sites derived from the model remained consistent across the different techniques used. The different wavelengths at which the measurements were made maintained the coherence of the results. [source] KATP channel blockade protects midbrain dopamine neurons by repressing a glia-to-neuron signaling cascade that ultimately disrupts mitochondrial calcium homeostasisJOURNAL OF NEUROCHEMISTRY, Issue 2 2010Damien Toulorge J. Neurochem. (2010) 114, 583,564. Abstract While KATP channels serve primarily as metabolic gatekeepers in excitable cells, they might also participate in other important cellular functions. Here, we demonstrate that KATP channel blockade with the sulfonylurea derivative glibenclamide provided robust protection to dopamine neurons undergoing spontaneous and selective degeneration in midbrain cultures. Unexpectedly, glibenclamide operated not by a direct effect on dopamine neurons but instead by halting the proliferation of a population of immature glial cells lacking astrocytic and microglial markers. The antimitotic effect of glibenclamide appeared essential to unmask a prosurvival phosphoinositide 3-kinase (PI3K)/Akt-dependent signaling pathway that controlled shuttling of calcium from endoplasmic reticulum to mitochondria in dopamine neurons. Preventing integrin-ligand interactions with a decoy ligand, the Arg-Gly-Asp-Ser sequence peptide, reproduced survival promotion by glibenclamide via a mechanism that also required PI3K/Akt-dependent regulation of mitochondrial calcium. Noticeably, Arg-Gly-Asp-Ser did not cause a reduction in glial cell numbers indicating that it prevented the death process downstream of the level at which glibenclamide intervenes. Based on these results, we propose that KATP channel blockade protected dopamine neurons by inhibiting a glia-to-neuron signaling pathway that propagates through integrin/ligand interactions and ultimately disrupts PI3K/Akt-dependent signaling and mitochondrial calcium homeostasis. [source] Fc, receptor I activation induces leukocyte recruitment and promotes aggravation of glomerulonephritis through the FcR, adaptorEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 4 2007Yutaka Kanamaru Abstract Myeloid cells bear Fc receptors (FcR) that mediate inflammatory signaling through the ITAM-containing FcR, adaptor. They express FcR,-associated Fc,RI, which modulate either activating or inhibitory signaling depending on the type of ligand interaction. The role of Fc,RI, in disease progression remains unknown, notably in IgA nephropathy (IgAN), one of major causes of end-stage renal disease, in which large amounts of circulating IgA-immune complexes (IC) may mediate receptor activation. To analyze the involvement of Fc,RI activation in glomerulonephritis (GN), we generated Tg mice expressing a mutated, signaling-incompetent, human Fc,RIR209L that cannot associate with FcR,. Like Fc,RIwt -Tg mice, they developed mesangial IgA deposits but not macrophage infiltration. Fc,RI activation in Fc,RIwt, but not in Fc,RIR209L, Tg mice resulted in marked inflammation with severe proteinuria and leukocyte infiltration in spontaneous IgAN or anti-glomerular basement membrane Ab-induced GN models. Receptor triggering of syngenically transferred Fc,RIwt Tg macrophages into non-Tg animals induced their recruitment into injured kidneys during GN development. Fc,RIwt cross-linking on macrophages activated MAP kinases and production of TNF-, and MCP-1. Moreover, IgA-IC from IgAN patients activated Fc,RI and induced TNF-, production. Thus, Fc,RI activation mediates GN progression by initiating a cytokine/chemokine cascade that promotes leukocyte recruitment and kidney damage. [source] Netrin-G2 and netrin-G2 ligand are both required for normal auditory responsivenessGENES, BRAIN AND BEHAVIOR, Issue 4 2008W. Zhang Mice in which netrin-G2 has been genetically inhibited do not startle to an acoustic stimulus, but otherwise perform normally through a behavioral test battery. Light microscopic examination of the inner ear showed no obvious structural abnormalities. Brainstem responses to acoustic stimuli (auditory brainstem responses, ABR) were also present, confirming the lack of any overarching defects in the inner ear or auditory nerve. Genetic inhibition of netrin-G2 ligand produced a nearly identical phenotype, that is, no startle with ABR present, and otherwise normal. This similarity confirms that these two proteins act in the same biological pathway. We have also determined that the affinity between the two proteins is strong, around 2.5 nm, similar to that observed between netrin-G1 and netrin-G1 ligand , 2.3 nm in our hands. The combination of equivalent phenotypes when genetically inhibited coupled with evidence of a strong biochemical interaction supports the notion of a receptor,ligand interaction between these two proteins in vivo. This interaction is critical for auditory synaptic responsiveness in the brain. [source] Surface T-cell antigen receptor expression and availability for long-term antigenic signalingIMMUNOLOGICAL REVIEWS, Issue 1 2003Adam G. Schrum Summary:, It is important to understand how T-cell antigen receptor (TCR) engagement and signaling are regulated throughout an immune response. This review examines the dynamics of surface TCR expression and signaling capacity during thymic and effector T-cell development. Although the TCR can undergo vast changes in surface expression, T cells remain capable of sustaining TCR engagement for long periods of time. This may be achieved by a combination of mechanisms that involve (a) controlling the quantity of surface TCR available for ligand interaction and (b) controlling the quality of surface TCR expression during T-cell activation. TCR signaling itself appears to be one of the main quantitative modulators of surface TCR expression, and it can cause both downregulation and upregulation at different times of T-cell activation. Recent studies indicate that the degree of upregulation is tunable by the strength of antigenic stimulation. There is evidence that qualitatively distinct forms of the TCR exist, and their potential role in sustained antigenic signaling is also discussed. A goal of future studies will be to better characterize these modulations in surface TCR expression and to clarify their impact on the regulation of immune responses. [source] Roles of proinflammatory cytokines and the Fas/Fas ligand interaction in the pathogenesis of inflammatory myopathiesIMMUNOLOGY, Issue 1pt2 2009Masahiro Kondo Summary Within the lesions of inflammatory myopathies, muscle fibres and invading mononuclear cells express Fas and Fas ligand (FasL), respectively. However, the roles of the Fas/FasL interaction in the pathogenesis of inflammatory myopathies are not fully understood. In the present study, we investigated the roles of proinflammatory cytokines and the Fas/FasL system in the pathogenesis of inflammatory myopathies. In vitro culturing of muscle cells with the proinflammatory cytokines interferon-,, tumour necrosis factor-,, and interleukin (IL)-1, synergistically increased Fas expression, susceptibility to Fas-mediated apoptosis, and the expression of cytoplasmic caspases 8 and 3. In addition, culturing of muscle cells with activated CD4+ T cells induced muscle cell apoptosis, which was partially inhibited by anti-FasL antibody. We also tested the possibility that T helper (Th) 17, which is an IL-17-producing helper T-cell subset that plays crucial roles in autoimmune and inflammatory responses, participates in the pathogenesis of inflammatory myopathies. Interestingly, in vitro culturing of dendritic cells with anti-Fas immunoglobulin M (IgM) or activated CD4+ T cells induced the expression of mRNA for IL-23p19, but not for IL-12p35, in addition to proinflammatory cytokines. Furthermore, IL-23p19 and IL-17 mRNAs were detected in the majority of biopsy samples from patients with inflammatory myopathies. Taken together, these results suggest that proinflammatory cytokines enhance Fas-mediated apoptosis of muscle cells, and that the Fas/FasL interaction between invading dendritic cells and CD4+ T cells induces local production of IL-23 and proinflammatory cytokines, which can promote the proliferation of Th17 cells and enhance Fas-mediated apoptosis of muscle cells, respectively. [source] Regulation of ligands for the activating receptor NKG2DIMMUNOLOGY, Issue 4 2007Anita R. Mistry Summary The outcome of an encounter between a cytotoxic cell and a potential target cell depends on the balance of signals from inhibitory and activating receptors. Natural Killer group 2D (NKG2D) has recently emerged as a major activating receptor on T lymphocytes and natural killer cells. In both humans and mice, multiple different genes encode ligands for NKG2D, and these ligands are non-classical major histocompatibility complex class I molecules. The NKG2D,ligand interaction triggers an activating signal in the cell expressing NKG2D and this promotes cytotoxic lysis of the cell expressing the ligand. Most normal tissues do not express ligands for NKG2D, but ligand expression has been documented in tumour and virus-infected cells, leading to lysis of these cells. Tight regulation of ligand expression is important. If there is inappropriate expression in normal tissues, this will favour autoimmune processes, whilst failure to up-regulate the ligands in pathological conditions would favour cancer development or dissemination of intracellular infection. [source] Enhanced Osteoclastogenesis in 4-1BB,Deficient Mice Caused by Reduced Interleukin-10,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 12 2006Hyun-Hee Shin PhD Abstract Enhanced osteoclastogenesis was observed in bone marrow,derived macrophage cells from 4-1BB,deficient mice than in those from wildtype mice. 4-1BB and 4-1BB ligand interaction may play a role at a certain stage of osteoclast formation through increased level of IL-10, a negative regulator of osteoclastogenesis. Introduction: 4-1BB is an inducible T-cell costimulatory molecule and a member of the TNF receptor family. The expression pattern of 4-1BB and 4-1BB ligand (4-1BBL) has suggested that 4-1BB plays a role not only in various responses related to innate immunity but also in bone metabolism. Materials and Methods: Osteoclast formation was evaluated in bone marrow,derived macrophage cells (BMMs) from wildtype and 4-1BB,deficient (4-1BB,/,) mice. Expression of interleukin-10 (IL-10) during osteoclast formation was analyzed at the mRNA and protein levels. Results: Expression of IL-10 was higher in RANKL-stimulated wildtype BMMs than 4-1BB,/, BMMs. When 4-1BBL was stimulated with 4-1BB,Fc fusion protein, the expression of IL-10 in BMMs increased. Neutralization of IL-10 was not as effective in preventing inhibition by IL-10 of osteoclast differentiation in 4-1BB,/, BMMs as in wildtype BMMs. When IL-10 was added to the culture medium, osteoclast formation was inhibited more efficiently in the 4-1BB,/, BMMs than in the wildtype BMMs. Conclusions: Interaction of 4-1BB and 4-1BBL stimulates IL-10 production through 4-1BBL signaling. 4-1BBL plays a role at a certain stage of osteoclast formation, and IL-10 may mediate this effect. The elevated level of osteoclastogenesis in 4-1BB,/, BMMs may thus be caused, in part, by a lower level of IL-10. [source] Characterization of site I of human serum albumin using spectroscopic analyses: Locational relations between regions Ib and Ic of site IJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 12 2004Keishi Yamasaki Abstract Site I of human serum albumin is an important and complex region for high-affinity binding of drugs. Equilibrium dialysis showed independent binding of dansyl- L -asparagine (DNSA) and n -alkyl p -aminobenzoates (p -ABEs) to regions Ib and Ic, respectively, in the pH range 6.0,9.0. However, individual binding of DNSA increased with pH in the same range. Binding of the four n -alkyl p -ABEs strongly perturbed the circular dichroism spectrum of bound DNSA, and the effect increased with concentration and the number of carbon atoms in the alkyl moiety. A similar effect was observed by increasing pH from 6.0 to 9.0, a pH range in which human serum albumin is known to undergo the neutral-to-base transition. The spectral changes propose spatial orientation changes of DNSA at region Ib. This proposal was supported by increased fluorescence anisotropy values: n -alkyl p -ABEs binding and the pH-dependent conformational change each restricted the mobility of the naphthalene ring of bound DNSA. Despite the similar effects on the spatial orientation of DNSA, clear differences were observed between the effects of n -alkyl p -ABEs and neutral-to-base transition. The former hardly changed the affinity and maximum fluorescence emission wavelength of bound DNSA; in contrast, the latter significantly affected them. The results give new information about site I and, according to our knowledge, represent a new type of ligand interaction, because the binding site of DNSA could be changed by simultaneous binding of the n -alkyl p -ABEs without affecting the binding constant. © 2004 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 93:3004,3012, 2004 [source] Protein thermal stability and phospholipid,protein interaction investigated by capillary isoelectric focusing with whole column imaging detectionJOURNAL OF SEPARATION SCIENCE, JSS, Issue 7 2006Tao Bo Abstract CIEF with whole column imaging detection (WCID) is an attractive technique for studying protein reaction and protein,ligand interaction due to its fast separation, simple operation, and high efficiency. In this study, two interesting applications by the CIEF-WCID were developed, involving the study of protein thermal stability and phospholipid,protein interaction. Four proteins (,-lactoglobulin B, trypsin inhibitor, phosphorylase b, and trypsinogen) with different pI, and two types of phospholipids, including phosphatidylcholine (PC) and phosphatidylserine (PS), were used for this purpose. First, the altered CIEF profiles of four proteins were exhibited due to conformational changes resulting from protein denaturation induced by a high incubation temperature at 60°C. It was demonstrated that the addition of a zwitterionic phospholipid (PC) played a crucial role in the thermal stability of targeted proteins, especially for trypsin inhibitor whose thermal stability was promoted with the addition of the PC vesicles at 60°C. Second, the zwitterionic (PC) and acidic (PS) phospholipids displayed completely different interactions with the proteins. The addition of PS vesicles modified the zwitterionic phospholipids to carry negative charges, which correspondingly changed the interaction between the phospholipid and the protein. Our study demonstrates that the CIEF-WCID is a powerful approach to study protein reaction and protein,ligand interaction with high efficiency, high selectivity, and fast separation. [source] Involvement of the ,3 E749ATSTFTN756 region in stabilizing integrin ,IIb,3 -ligand interactionJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 10 2003P. E. M. H. Litjens Summary., Platelet integrin ,IIb,3 must be activated via intracellular mechanisms before it binds soluble ligands, and it is thought to be activated at its extracellular site by surface-bound ligands. Integrin activation is associated with rearrangement of the cytoskeleton and phosphorylation of proteins that become localized in focal contacts. In these processes, the cytoplasmic tail of the ,-subunit plays a central role. We introduced peptides homologous to the E749ATSTFTN756 domain (E,N peptide) and the T755NITYRGT762 domain (T,T peptide) of ,3 in streptolysin O-permeabilized platelets and analyzed the initial interaction with soluble fibronectin, fibrinogen and PAC-1 after stimulation with thrombin. E,N peptide left the initial binding of fibronectin intact but interfered with stable receptor occupancy. E,N peptide also inhibited fibrinogen binding, thereby reducing the formation of large aggregates. Strikingly, E,N peptide did not disturb the binding of PAC-1, which is known to reflect activation of the integrin. E,N peptide also inhibited tyrosine phosphorylation of focal adhesion kinase, a response known to be dependent on ,IIb,3. T,T peptide did not affect these processes. In a model for outside-in integrin activation, E,N peptide disrupted the binding of CHO cells expressing ,IIb,3 to surface-bound ligand. Again, T,T peptide had no effect. We conclude that the E749ATSTFTN756 region of the ,3 -tail stabilizes the binding of soluble and surface-bound ligand to integrin ,IIb,3 via a mechanism that involves the phosphorylation of FAK. [source] CC Chemokine Receptor 4 (CCR4) in human allergen-induced late nasal responsesALLERGY, Issue 9 2010G. Banfield To cite this article: Banfield G, Watanabe H, Scadding G, Jacobson MR, Till SJ, Hall DA, Robinson DS, Lloyd CM, Nouri-Aria KT, Durham SR. CC Chemokine Receptor 4 (CCR4) in human allergen-induced late nasal responses. Allergy 2010; 65: 1126,1133. Abstract Background:, CC Chemokine receptor 4 (CCR4) is preferentially expressed on Th2 lymphocytes. CCR4-mediated inflammation may be important in the pathology of allergic rhinitis. Disruption of CCR4 , ligand interaction may abrogate allergen-induced inflammation. Methods:, Sixteen allergic rhinitics and six nonatopic individuals underwent both allergen and control (diluent) nasal challenges. Symptom scores and peak nasal inspiratory flow were recorded. Nasal biopsies were taken at 8 h post challenge. Sections were immunostained and examined by light or dual immunofluorescence microscopy for eosinophils, T-lymphocytes, CCR4+CD3+ and CXCR3+CD3+ cells and examined by in situ hybridization for CCR4, IL-4 and IFN-, mRNA+ cells. Peripheral blood mononuclear cells were obtained from peripheral blood of nine normal donors and the CCR4+CD4+ cells assessed for actin polymerization in response to the CCR4 ligand macrophage-derived chemokine (MDC/CCL22) and the influence of a CCR4 antagonist tested. Results:, Allergic rhinitics had increased early and late phase symptoms after allergen challenge compared to diluent; nonatopics did not respond to either challenge. Eosinophils, but not total numbers of CD3+ T cells, were increased in rhinitics following allergen challenge. In rhinitics, there was an increase in CCR4+CD3+ protein-positive cells relative to CXCR3+CD3+ cells; CCR4 mRNA+ cells were increased and IL-4 increased to a greater extent than IFN-,. CCR4+CD4+ T cells responded to MDC in vitro, and this response was inhibited by the selective CCR4 antagonist. Conclusion:, Lymphocyte CCR4 expression is closely associated with induction of human allergen-induced late nasal responses. Blocking CCR4-ligand interaction may provide a novel therapeutic approach in allergic disease. [source] Getting down to malarial nuts and bolts: the interaction between Plasmodium vivax merozoites and their host erythrocytesMOLECULAR MICROBIOLOGY, Issue 5 2005Julian Rayner Summary Of the four Plasmodium species that routinely cause malaria in humans, Plasmodium falciparum is responsible for the majority of malaria mortality and consequently gets most of the headlines. Outside Africa, however, more malaria cases are caused by its distant cousin Plasmodium vivax, resulting in a daunting morbidity and economic burden for countries across Asia and the Americas. Plasmodium life cycles are complex, but the symptoms and pathology of malaria occur during the blood phase, when merozoites recognize and invade erythrocytes, initiating a developmental programme that culminates in lysis of the erythrocyte and release of multiple daughter merozoites. P. vivax merozoites are dependent on a single host cell receptor for erythrocyte invasion, the Duffy antigen receptor for chemokines, and humans that do not express this receptor on the surface of their erythrocytes are immune to P. vivax infection. This essential receptor,ligand interaction is addressed from both the host and parasite side in two papers in this issue of Molecular Microbiology, with important implications for plans to develop a P. vivax vaccine. [source] Elucidation of the mechanism of the regulatory function of the Ig1 module of the fibroblast growth factor receptor 1PROTEIN SCIENCE, Issue 10 2006Vladislav V. Kiselyov Abstract The extracellular part of the fibroblast growth factor (FGF) receptor (FGFR) consists of up to three Ig modules (Ig1,Ig3), in which the Ig2 and Ig3 modules determine affinity and specificity for FGF and heparin. The FGFR isoforms lacking the Ig1 module have higher affinity for FGF and heparin than the triple Ig-module isoforms, suggesting that the Ig1 module is involved in the regulation of the FGFR,ligand interaction. We show here by surface plasmon resonance and NMR analyses that the Ig1 module binds to the Ig2 module, and identify by NMR the binding sites involved in the Ig1,Ig2 interaction. The identified binding site in the Ig2 module was found to be in the area of the FGF,Ig2 and Ig2,heparin contact sites, thus providing direct structural evidence that the Ig1 module functions as a competitive autoinhibitor of the FGFR,ligand interaction. Furthermore, the Ig1 binding site of the Ig2 module overlaps the Ig2,Ig2 contact site. This suggests that the function of the Ig1 module is not only regulation of the FGFR,ligand binding affinity but also prevention of spontaneous FGFR dimerization (through a direct Ig2,Ig2 interaction) in the absence of FGF. [source] Fas-mediated upregulation of vascular endothelial growth factor and monocyte chemoattractant protein-1 expression in cultured dermal fibroblasts: Role in the inflammatory responseTHE JOURNAL OF DERMATOLOGY, Issue 2 2007Masao FUJIWARA ABSTRACT The Fas,Fas ligand interaction is the most important pathway in starting apoptosis. In addition, several recent reports have emerged documenting non-apoptotic roles for Fas. However, a non-apoptotic role of Fas in dermal fibroblasts remains unknown. The present study investigated whether Fas stimulation not only promotes apoptosis but also stimulates elements of the inflammatory response such as angiogenesis and macrophage infiltration. Fas stimulation was performed by treating cultured human dermal fibroblasts with an agonistic anti-Fas monoclonal antibody (mAb). Anti-Fas mAb-treated fibroblasts showed a significantly greater increase of caspase-3 and caspase-8 activity compared with control fibroblasts. Addition of the anti-Fas mAb induced DNA fragmentation, as confirmed by the DNA ladder assay. Terminal deoxynucleotidyl transferase-mediated 2,-deoxyuridine 5,-triphosphate nick end labeling (TUNEL) staining showed that treatment with the anti-Fas mAb induced an increase of apoptotic fibroblasts in a time-dependent manner. At both mRNA and protein levels, anti-Fas mAb-treated fibroblasts showed significantly higher expression of vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein (MCP)-1 compared with control fibroblasts. A pan-caspase inhibitor (Z-VAD-FMK) significantly inhibited VEGF and MCP-1 expression. After transplantation of fibroblasts into mice with severe combined immunodeficiency, the nodules derived from anti-Fas mAb-treated fibroblasts showed more abundant neovascularization, increased macrophage infiltration, and more apoptotic cells in comparison with nodules derived from control fibroblasts. The results of both in vitro and in vivo studies confirmed significantly higher angiogenic activity and macrophage chemotactic activity of anti-Fas mAb-treated fibroblasts compared with control fibroblasts. [source] Life-science applications of the Cambridge Structural DatabaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 6-1 2002Robin Taylor Several studies show that the molecular geometries and intermolecular interactions observed in small-molecule crystal structures are relevant to the modelling of in vivo situations, although the influence of crystal packing is sometimes important and should always be borne in mind. Torsional distributions derived from the Cambridge Structural Database (CSD) can be used to map out potential-energy surfaces and thereby help identify experimentally validated conformational minima of molecules with several rotatable bonds. The use of crystallographic data in this way is complementary to in vacuo theoretical calculations since it gives insights into conformational preferences in condensed-phase situations. Crystallographic data also underpin many molecular-fragment libraries and programs for generating three-dimensional models from two-dimensional chemical structures. The modelling of ligand binding to metalloenzymes is assisted by information in the CSD on preferred coordination numbers and geometries. CSD data on intermolecular interactions are useful in structure-based inhibitor design both in indicating how probable a protein,ligand interaction is and what its geometry is likely to be. They can also be used to guide searches for bioisosteric replacements. Crystallographically derived information has contributed to many life-science software applications, including programs for locating binding `hot spots' on proteins, docking ligands into enzyme active sites, de novo ligand design, molecular superposition and three-dimensional QSAR. Overall, crystallographic data in general, and the CSD in particular, are very significant tools for the rational design of biologically active molecules. [source] Prenylflavonoids as Nonsteroidal Phytoestrogens and Related Structure,Activity RelationshipsCHEMMEDCHEM, Issue 4 2006Zhi-qiang Wang Abstract In the search for estrogen receptor (ER) modulators, a series of prenylflavonoids were found to be widely distributed amongst tonic herbal medicines and to possess estrogen-like activity in MCF-7/BOS cells, as evaluated by an estrogen-screening assay. Cell-cycle analysis revealed that the stimulatory effects of these compounds toward cell proliferation were elicited at the G1,S checkpoint and could significantly increase the S-phase population of MCF-7 cells under hormone-free conditions. ER-responsive gene (PS2, PgR) and protein (PgR) expression was also detected; mRNA and protein-expression levels for PS2 and PgR were up-regulated by the compounds in a dose-dependent manner. These effects could be inhibited by the pure ER antagonist ICI,182,780 ((7,-[9-{4,4,5,5,5-pentafluoropentyl}sulfinyl]nonyl)estra-1,3,5(10)-triene-3,17,-diol). It was therefore concluded that the estrogen-like effects of these prenylflavonoids were mediated primarily through ERs. Furthermore, to explore the structure,activity relationship based on the estrogen receptor and detailed molecular mechanisms among the prenylflavonoids, protein,ligand docking simulations were carried out by using the DS-MODELING software package. The binding affinity of each prenylflavonoid toward ER, was scored, and the receptor,ligand interaction was also analyzed to provide the simulation characteristics of virtual molecular recognition mechanisms. [source] Two-Dimensional Self-Assembly of a Porphyrin,Polypyridyl Ruthenium(II) Hybrid on HOPG Surface through Metal,Ligand InteractionsCHEMPHYSCHEM, Issue 9 2010Aimei Gao Dr. Abstract The synthesis and self-assembly behavior of porphyrin,polypyridyl ruthenium(II) hybrid, which consists of a flexible alkyl chain attached with two conjugated moieties is described. The electronic absorption spectrum and emission spectra show that the [C8 -TPP-(ip)Ru(phen)2](ClO4)2, abbreviated as (C8ip)TPPC has optical properties. Scanning tunneling microscopy (STM) studies found that the ,,, interaction and metal,ligand interaction allow (C8ip)TPPC to form self-assembled structure and have an edge-on orientation on the highly oriented pyrolytic graphite (HOPG) surface. The multidentate structure in (C8ip)TPPC molecules act as linkers between the molecules and form metal,ligand coordination, which forces the assembly process in the direction of stable columnar arrays. In addition, although the sample was stored for two months in ambient conditions, STM experiments showed that the order of (C8ip)TPPC self-assembly only slightly decreased which indicates that the self-assembled monolayer is stable. This work demonstrates that introducing a metal-ligand in the porphyrin-polypyridyl compound is a useful strategy to obtain novel surface assemblies. [source] CE-ESI-MS/MS as a rapid screening tool for the comparison of protein,ligand interactionsELECTROPHORESIS, Issue 7 2010Thomas Hoffmann Abstract In drug development, the combinatorial synthesis of drug libraries is common use, therefore efficient tools for the characterization of drug candidates and the extent of interaction between a drug and its target protein is a central question of analytical interest. While biological activity is tested today by enzyme assays, MS techniques attract more and more attention as an alternative for a rapid comparison of drug,target interactions. CE enables the separation of proteins and drug,enzyme complexes preserving their physiological activity in aqueous media. By hyphenating CE with ESI-MS/MS, the binding strength of enzyme inhibitors can be deduced from MS/MS experiments, which selectively release the inhibitor from the drug,target complex after CID. In this study, ,-chymotrypsin (CT), a serine protease, was chosen as a model compound. Chymostatin is a naturally occurring peptide aldehyde binding to CT through a hemiacetal bond and electrostatic interaction. First, a CE separation was developed, which allows the analysis of ,-CT and a chymotrypsin,chymostatin complex under MS-compatible conditions. The use of neutral-coated CE capillaries was mandatory to reduce analyte,wall interactions. ESI-quadrupole ion trap-MS was worked out to demonstrate the selective drug release after CID. Fragmentation of the drug,enzyme complex was monitored in dependence from the excitation energy in the ion trap, leading to the V50 voltage that enables 50% complex fragmentation as a reference value for chymotrypsin,chymostatin complex. A stable CE-ESI-MS/MS setup was established, which preserves the drug,enzyme complexes during ionization,desolvation processes. With this optimized setup, different CT inhibitors could be investigated and compared. [source] Prevention of diabetes in NOD mice at a late stage by targeting OX40/OX40 ligand interactionsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 11 2004Syamasundar Abstract Autoreactive T,cells play a major role in the development of insulin-dependent diabetes mellitus, suggesting that costimulatory molecules that regulate T,cell responses might be essential for disease progression. In NOD mice, CD28/B7 and CD40/CD40 ligand,(L) interactions control the onset of diabetes from 2 to 4,weeks of age, but blocking these molecules has little effect after this time. Hence, it is possible that other ligand/receptor pairs control a later phase of disease. We now show that OX40 is expressed on CD4 and CD8 T,cells several weeks prior to islet destruction, which is initiated around weeks,12,14, and that OX40L is present on dendritic cells in both secondary lymphoid organs and the pancreas from 11 to 13,weeks of age. Blocking OX40L at 6, 9, or 15,weeks after birth had little effect on disease; however, inhibiting OX40/OX40L interactions at week,12, or continuous treatment from week,12 onwards, significantly reduced the incidence of diabetes. Histological examination showed that islet destruction was prevented and insulitis reduced by targeting OX40L. These studies show that OX40/OX40L interactions form a late checkpoint in diabetes development and suggest that these molecules are realistic targets for therapeutic intervention. [source] Fine-Tuning Ligands for Catalysis Using Supramolecular StrategiesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 29 2007Vincent F. Slagt Abstract Coordinative bonds have been used to prepare supramolecular ligands leading to well-defined catalysts formed by assembly. The construction of these ligands is based on selective metal,ligand interactions between nitrogen donor atoms of phosphorus-nitrogen building blocks and various zinc(II) porphyrins. The major advantage of this supramolecular approach of catalyst preparation is the simplification of ligand variation enabling straightforward modification of steric, electronic and chiral properties of the supramolecular ligand. A large number of new ligands becomes accessible by this modular variation of the building blocks. The ligand assembly based on pyridyl phosphites and zinc(II) porphyrin with electron-withdrawing substituents led to a twelve-fold increase in activity and an increase in enantioselectivity from 17 to 50,% in the rhodium-catalyzed hydrogenation of dimethyl itaconate. The first examples of assemblies based on non-chiral ligands and chiral zinc(II) porphyrin template molecules show, as proof of principle, an enantiomeric excess up to 18,% in the asymmetric palladium-catalyzed allylic alkylation. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Comparison of the inward- and outward-open homology models and ligand binding of human P-glycoproteinFEBS JOURNAL, Issue 23 2009Ilza K. Pajeva An homology model of human P-glycoprotein, based on the X-ray structure of the recently resolved mouse P-glycoprotein, is presented. The model corresponds to the inward-facing conformation competent for drug binding. From the model, the residues involved in the protein-binding cavity are identified and compared with those in the outward-facing conformation of human P-glycoprotein developed previously based on the Sav1866 structure. A detailed analysis of the interactions of the cyclic peptides QZ59- RRR and QZ59- SSS is presented in both the X-ray structures of mouse P-glycoprotein and the human P-glycoprotein model generated by ligand docking. The results confirm the functional role of transmembrane domains TM4, TM6, TM10 and TM12 as entrance gates to the protein cavity, and also imply differences in their functions. The analysis of the cavities in both models suggests that the ligands remain bound to the same residues during the transition from the inward- to the outward-facing conformations. The analysis of the ligand,protein interactions in the X-ray complexes shows differences in the residues involved, as well as in the specific interactions performed by the same ligand within the same protein. This observation is supported by docking of the QZ59 ligands into human P-glycoprotein, thus aiding in the understanding of the complex behavior of P-glycoprotein substrates and inhibitors. The results confirm the possibility for multispecific drug interactions of the protein, and are important for elucidating the P-glycoprotein function and ligand interactions. [source] Regulation of signal transduction by glycosylationINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 4 2004Robert S. Haltiwanger The incredible diversity found in cell-surface glycoconjugate structures led researchers over 30 years ago to propose that complexity in carbohydrates must play a role in cellular communication. Recent studies from a number of laboratories have confirmed this hypothesis, demonstrating that cell-surface glycoconjugates play significant roles in signal transduction events. One striking example is the effect of O -fucose modifications on the Notch-signalling pathway. Notch is a cell-surface receptor that is essential for proper development. The extracellular domain of Notch contains up to 36-tandem epidermal growth factor-like (EGF) repeats, many of which are predicted to be modified at putative consensus sequences with O -fucose and O -glucose saccharides. Genetic alterations (by knockout or RNAi methodologies) in the enzyme responsible for the addition of O -fucose to Notch, protein O -fucosyltransferase-1, result in severe, embryonic lethal phenotypes resembling Notch mutants. Thus, O -fucosylation appears to be essential for proper Notch function. Elongation of the O -fucose monosaccharide by the ,1,3- N -acetylglucosaminyltransferase, Fringe, modulates Notch function, either increasing or decreasing response from ligands depending on context. Although it is now clear that O -fucose modifications affect Notch signalling, the molecular mechanism by which this occurs is not known. As an initial step in understanding how O -fucose glycans affect Notch function, we are mapping O -fucose modifications to specific sites on Notch. Already, we have demonstrated that O -fucose modifies one of the EGF repeats involved in ligand binding, suggesting that the sugars may play a role in Notch,ligand interactions. Experiments to test the role of O -fucose modifications at specific sites are in progress. We have also found that Fringe modifies O -fucose on some EGF repeats but not others. Our initial analyses suggest that the basis of this specificity is encoded within the sequences of the individual EGF repeats. Site mapping has also confirmed the presence of O -glucose saccharides on Notch. The evolutionarily conserved, predicted O -glucose sites on Notch are as numerous as those for O -fucose, suggesting that the O -glucose modifications will play an equally important role in Notch biology. We have recently identified an enzymatic activity capable of catalyzing the addition of O -glucose to EGF repeats. Purification of the protein O -glucosyltransferase is underway. These and other results will be discussed. [source] Use of [125I]4,-iodoflavone as a tool to characterize ligand-dependent differences in Ah receptor behaviorJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 6 2002Hollie I. Swanson Abstract We have synthesized [125I]4,-iodoflavone to study Ah receptor (AhR),ligand interactions by a class of AhR ligands distinct from the prototypic ligand 2,3,7,8-tetrachlorodibenzo- p -dioxin (TCDD). This radioligand allows the comparison of AhR,ligand interactions using a ligand that differs in AhR affinity, and yet has the same radiospecific activity as [125I]2-iodo-7,8-dibromodibenzo- p -dioxin. Specific binding of [125I]4,-iodoflavone with the AhR was detected as a single radioactive peak (,9.7 S) following density sucrose gradient analysis. Cytosolic extracts from both Hepa 1 and HeLa cells were used as the source of mouse and human AhR, respectively. A ,6.7 S form of radioligand-bound Ah receptor was detected in the high salt nuclear extracts of both cell lines. In HeLa cells approximately twofold more [125I]4,-iodoflavone,AhR 6 S complex, compared with [125I]2-iodo-7,8-dibromodibenzo- p -dioxin, was recovered in nuclear extracts. A comparison of the ability of 4,-iodoflavone and TCDD to cause time-dependent translocation of AhR-yellow fluorescent protein revealed that 4,-iodoflavone was more efficient at enhancing nuclear accumulation of the receptor. These results suggest that [125I]4,-iodoflavone is a particularly useful and easily synthesized ligand for studying the AhR. © 2002 Wiley Periodicals, Inc. J Biochem Mol Toxicol 16:298,310, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.10053 [source] | ||||||||||||||||||||||||||||||||||