Corticotropin releasing factor (CRF) receptor antagonists are under preclinical and clinical investigation for stress-related disorders. In this study the impact of receptor-ligand binding kinetics on CRF1 receptor antagonist pharmacology was investigated by measuring the association rate constant (k1), dissociation rate constant (k-1) and kinetically-derived affinity at 37°C. Three aspects of ...

molA (HI1472) from H. influenzae encodes a periplasmic binding protein (PBP) that delivers substrate to the ABC transporter MolB(2)C(2) (formerly HI1470/71). The structures of MolA with molybdate and tungstate in the binding pocket were solved to 1.6 and 1.7 Å resolution, respectively. The MolA-binding protein binds molybdate and tungstate, but ...

The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family plays a central role in antigenic variation and cytoadhesion of P. falciparum infected erythrocytes. PfEMP1 proteins/var genes are classified into three main subfamilies (UpsA, UpsB, and UpsC) that are hypothesized to have different roles in binding and disease. To investigate whether ...

The type II secretion system (T2SS) is a large macromolecular complex spanning the inner and outer membranes of many gram-negative bacteria. The T2SS is responsible for the secretion of virulence factors such as cholera toxin (CT) and heat-labile enterotoxin (LT) from Vibrio cholerae and enterotoxigenic Escherichia coli, respectively. CT and ...

Recently, a new model for glutamate uptake by glutamate transporters was proposed based on crystal structures of the bacterial glutamate transporter homolog Glt(Ph). It was proposed that hairpin two (HP2) functions as the extracellular gate and that Na(+) and glutamate binding closes HP2, thereby allowing for the translocation of the ...

Herpes simplex virus (HSV) entry requires the core fusion machinery of gH/gL and gB as well as gD and a gD receptor. When gD binds receptor, it undergoes conformational changes that presumably activate gH/gL, which then activates gB to carry out fusion. gB is a class III viral fusion protein, ...

In this study, we used potentiometric titrations and Cd adsorption experiments to determine the binding capacities of two acidophilic (A. cryptum and A. acidophilum) and two alkaliphilic (B. pseudofirmus and B. circulans) bacterial species in order to determine if any consistent trends could be observed relating bacterial growth environment to ...

The lac repressor is an allosteric protein that can respond to environmental changes. Mutationsintroduced into the DNA binding domain and the effector binding pocketaffect the repressor's ability to respond to its environment. We have demonstrated how the observed phenotype is a consequence of altering the thermodynamic equilibrium constants. Mutant repressors ...

Two chromogenic complexes, L.Zn (where L is (E)-4-((4-(1,4,8,11-tetraazacyclotetradecan-1-ylsulfonyl)phenyl)diazenyl)-N,N-dimethylaniline) and its [2]pseudorotaxane form (α-CD.L.Zn), were found to bind preferentially to adenosine triphosphate (ATP), among all other common anions and biologically important phosphate (AMP, ADP, pyrophosphate, and phosphate) ions in aqueous HEPES buffer medium of pH 7.2. Studies with live cell cultures ...

Four-phosphate-adaptor protein 1 (FAPP1) regulates secretory transport from the trans-Golgi network (TGN) to the plasma membrane. FAPP1 is recruited to the Golgi through binding of its pleckstrin homology (PH) domain to phosphatidylinositol 4-phosphate (PtdIns(4)P) and a small GTPase ADP-ribosylation factor 1 (ARF1). Despite the critical role of FAPP1 in membrane ...

The neurofibromatosis type 2 tumor suppressor protein, merlin, is related to the ERM (ezrin, radixin, and moesin) family of plasma membrane-actin cytoskeleton linkers. For ezrin, phosphatidylinositol 4,5-bisphosphate (PIP(2)) binding to the amino-terminal FERM domain is required for its conformational activation, proper subcellular localization, and function, but less is known about ...

Although membrane proteins are ubiquitous within all living organisms and represent the majority of drug targets, a general method for direct, label-free measurement of ligand binding to native membranes has not been reported. Here we show that backscattering interferometry (BSI) can accurately quantify ligand-receptor binding affinities in a variety of ...

The Clostridium botulinum neurotoxins (BoNTs) cleave SNARE proteins, which inhibit binding and thus fusion of neurotransmitter vesicles to the plasma membrane of peripheral neurons. BoNTs comprise an N-terminal light chain (LC) and C-terminal heavy chain, which are linked by a disulfide bond. There are seven serotypes (A-G) of BoNTs based ...

Replication of the bacterial chromosome is initiated by the binding of the DnaA protein to oriC. Various factors control the ability of DnaA to bind and unwind DNA. Among them E. coli DiaA and H. pylori HobA have been recently characterized. They interact with domain I of DnaA and stimulate ...

Proteinaceous stalks produced by Gram-negative bacteria are often used to adhere to environmental surfaces. Among them, chaperone-usher (CU) fimbriae adhesins, related to prototypical type 1 fimbriae, interact in highly specific ways with different ligands at different stages of bacterial infection or surface colonisation. Recent analyses revealed a large number of ...

ITC titration studies of a family of bis-ammonium receptors based upon a scaffold of two bis-linked phenol rings show that several of the receptors bind to both dihydrogenphosphate and phosphatidylglycerol anions in a similar binding motif. Thermodynamic properties determined from ITC show that anion binding is entropy driven. Job plots ...

Multivalency is playing a major role in biological processes and particularly in lectin-carbohydrate interactions. The design of high-affinity ligands of lectins should provide molecules capable of interfering with these biological processes and potentially inhibit bacterial or viral infections. Azide-alkyne "click" chemistry was applied to the synthesis of dodecavalent fullerene-based glycoclusters. ...

Interactions between fibronectin and tenascin-C within the extracellular matrix provide specific environmental cues that dictate tissue structure and cell function. The major binding site for fibronectin lies within the fibronectin type III-like repeats (TNfn) of tenascin-C. Here, we systematically screened TNfn domains for their ability to bind to both soluble ...

Sulfur is an essential element for microorganisms and it can be obtained from varied compounds, sulfate being the preferred source. The first step for sulfate assimilation, sulfate uptake, has been studied in several bacterial species. This article reviews the properties of different bacterial (and archaeal) transporters for sulfate, molybdate, and ...

Bacterial pathogens secrete effectors into their hosts that subvert host defenses and redirect host processes. EspG is a type three secretion effector with a disputed function that is found in enteropathogenic Escherichia coli. Here we show that EspG is structurally similar to VirA, a Shigella virulence factor; EspG has a ...

A gram-negative Sphingomonas sp. A1 directly incorporates alginate polysaccharide into the cytoplasm via the cell-surface pit and ABC transporter. A cell-surface alginate-binding protein, Algp7, functions as a concentrator of the polysaccharide in the pit. Based on the primary structure and genetic organization in the bacterial genome, Algp7 was found to ...

Candida albicans binds and utilizes human complement inhibitors, such as C4b-binding protein (C4BP), Factor H, and FHL-1 for immune evasion. Here, we identify Candida pH-regulated antigen 1 (Pra1) as the first fungal C4BP-binding protein. Recombinant Pra1 binds C4BP, as shown by ELISA and isothermal titration calorimetry, and the Pra1-C4BP interaction ...

Proteins use conserved binding motifs associated with relatively unconserved flexible amino acid sequences as mobile tethers for interacting molecules, as exemplified by C-terminal sequences of bacterial chemotaxis receptors. The FLEXTAIL bioinformatics pipeline predicts flexible tethers and their binding motifs based on the properties of flexibility and sequence conservation. In four ...

Bacterial luciferase contains an extended 29-residue mobile loop. Movements of this loop are governed by binding of either flavin mononucleotide (FMNH2) or polyvalent anions. To understand this process, loop dynamics were investigated using replica-exchange molecular dynamics that yielded conformational ensembles in either the presence or absence of FMNH2. The resulting ...

We have used single-walled carbon nanotube field-effect transistor (NTFET) devices to probe the interactions between carbohydrates and their recognition proteins called lectins. These interactions are involved in a wide range of biological processes, such as cell-cell recognition, cell-matrix interaction as well as viral and bacterial infections. In our experiments, NTFETs ...

Polymyxin peptide conjugated to the end groups of highly branched poly(N-isopropyl acrylamide) was shown to bind to a Gram negative bacterium, Pseudomonas aeruginosa . The nonbound polymer had a lower critical solution temperature (LCST) above 60 °C. However, binding caused aggregation, which was disrupted on cooling of the bacteria and ...

The conformational change observed upon ligand binding and phosphorylation for the cAMP-dependent protein kinase (protein kinase A-PKA) is of high importance for the regulation of its activity. We calculate pKa values and net charges for 18 3D structures of PKA in various conformations and liganded states to examine the role ...

Biocompatible glyco-dendronized poly-l-lysine (PLL) polymers carry either three or nine mannose- or galactose-bearing dendrons that selectively bind, and thus can be used to detect, bacteria. Central to the synthesis of glyco-dendronized polymers was the development of a continuous flow [2 + 2] photocycloaddition reaction to connect the dendrons and PLL. ...

Langerin mediates the carbohydrate-dependent uptake of pathogens by Langerhans cells in the first step of antigen presentation to the adaptive immune system. Langerin binds to an unusually diverse number of endogenous and pathogenic cell surface carbohydrates, including mannose-containing O-specific polysaccharides derived from bacterial lipopolysaccharides identified here by probing a microarray ...

The porins OmpF and OmpC are trimeric β-barrel proteins with narrow channels running through each monomer that exclude molecules > 600 Da while mediating the passive diffusion of small nutrients and metabolites across the Gram-negative outer membrane (OM). Here, we elucidate the mechanism by which an entire soluble protein domain ...

The pH-dependent binding affinity of either avidin or streptavidin for iminobiotin has been utilized in studies ranging from affinity binding chromatography to dynamic force spectroscopy. Regardless of which protein is used, the logarithmic dependence of the equilibrium dissociation constant (K(d)) on pH is assumed conserved. However a discrepancy has emerged ...

In this work, we investigated the feasibility of using phenyl boronate (PB) chromatography for the direct capture of monoclonal antibodies from a CHO cell supernatant. Preliminary results, using pure protein solutions have shown that PB media can bind to human antibodies, not only at strong alkaline conditions but also at ...

UNC-104/KIF1A is a Kinesin-3 motor that transports synaptic vesicles from the cell body towards the synapse by binding to PI(4,5)P(2) through its PH domain. The fate of the motor upon reaching the synapse is not known. We found that wild-type UNC-104 is degraded at synaptic regions through the ubiquitin pathway ...

In this letter, a pH-switchable system for protein adsorption and release is introduced. By combining the pH sensitivity of poly(methacrylic acid) (poly(MAA) chains and the nanoeffects of 3D nanostructured silicon nanowire arrays (SiNWAs), a poly(MAA)-modified SiNWAs material showed an extremely high capacity for binding lysozyme at pH 4 (an ∼80-fold ...

The complexation of arsenic (As) with dissolved organic matter (DOM), although playing an important role in regulating As mobility and transformation, is poorly characterized, as evidenced by scarce reporting of fundamental parameters of As-DOM complexes. The complexation of arsenite (AsIII) with Aldrich humic acid (HA) at different pHs was characterized ...

The PI3-kinase (PI3K) pathway regulates many cellular processes, especially cell metabolism, cell survival, and apoptosis. Phosphatidylinositol-3,4,5-trisphosphate (PIP3), the product of PI3K activity and a key signaling molecule, acts by recruiting pleckstrin-homology (PH) domain-containing proteins to cell membranes. Here, we describe a new structural class of nonphosphoinositide small molecule antagonists (PITenins, ...

Ferredoxin:NADPH oxidoreductase (FNR) is a key enzyme of photosynthetic electron transport required for generation of reduction equivalents. Recently, two proteins were found to be involved in membrane-anchoring of FNR by specific interaction via a conserved Ser/Pro-rich motif: Tic62 and Trol. Our crystallographic study reveals that the FNR-binding motif, which forms ...

Potentiators are molecules that increase the activity of the cystic fibrosis transmembrane conductance regulator (CFTR). Some potentiators can also inhibit CFTR at higher concentrations. The activating binding site is thought to be located at the interface of the dimer formed by the two nucleotide-binding domains. We have hypothesized that if ...

A new binding agent, consisting of the yeast Saccharomyces cerevisiae immobilized in agarose, is proposed for use in diffusive gradients in thin films (DGT). Different gel compositions, containing from 4.5% to 20% (m/v) of S. cerevisiae and 1.5-5.0% (m/v) of agarose, were prepared and tested for uptake of Cd(II). For ...

Culex mosquitoes introduce the pathogens responsible for filariasis, West Nile virus, St. Louis encephalitis, and other diseases into humans. Currently, traps baited with oviposition semiochemicals play an important role in detection efforts and could provide an environmentally friendly approach to controlling their populations. The odorant binding proteins (OBPs) in the ...

Binding of antibiotics to clay minerals can decrease both their physical and biological availability in soils. To elucidate the binding mechanisms of tetracycline antibiotics on smectite clays as a function of pH, we probed the interactions of oxytetracycline (OTC) with Na-montmorillonite (MONT) using X-ray diffraction (XRD), infrared (IR), and solid-state ...

Measurements at high spatial resolution by DGT (diffusive gradients in thin films) require a binding agent that is homogeneously distributed in the binding layer. Formation of ferrihydrite by in situ precipitation within a hydrogel has been previously shown to meet these requirements for the measurement of oxyanions by DGT. Here, ...

Clay-humic substance complexes play a major role in controlling the mobility of toxic metals in contaminated soils. However, our understanding of the underlying mechanisms is limited. Binding site analysis of clay and clay-mineral-humic composites, in this study, revealed an enhanced surface reactivity for the clay surface by the sorbed humic ...

A strategy was developed to produce thin, biopolymer-based polyelectrolyte multilayer films, based on hyaluronic acid and chitosan, that are able to effectively bind B lymphocytes. These films explore CD44-hyaluronate interactions and provide a method to make surface-bound B cell arrays without the need for nonselective covalent chemistry. The rational design ...

Anti-TRAP (AT) is a small zinc-binding protein that regulates tryptophan biosynthesis in Bacillus subtilis by binding to tryptophan-bound trp RNA-binding attenuation protein (TRAP), thereby preventing it from binding RNA, and allowing transcription and translation of the trpEDCFBA operon. Crystallographic and sedimentation studies have shown that AT can homooligomerize to form ...

A competitive-ligand, equilibrium-dialysis technique using bromide measured methylmercury (MeHg(+)) binding to Suwannee River fulvic acid (SRFA) and NOM from a lake and a bog in Minnesota. Distribution coefficients (K(OC)) and stability constants (K') varied only slightly over a range of [Br(-)] and ratios of MeHg(+) to reduced sulfur, S(re), the ...

The large GTPase dynamin has an important membrane scission function in receptor-mediated endocytosis and other cellular processes. Self-assembly on phosphoinositide-containing membranes stimulates dynamin GTPase activity, which is crucial for its function. Although the pleckstrin-homology (PH) domain is known to mediate phosphoinositide binding by dynamin, it remains unclear how this promotes ...

Helicobacter pylori , a pathogen that colonizes the human stomach, requires the nickel-containing metalloenzymes urease and NiFe-hydrogenase to survive this low pH environment. The maturation of both enzymes depends on the metallochaperone, HypA. HypA contains two metal sites, an intrinsic zinc site and a low-affinity nickel binding site. X-ray absorption ...

To characterize the specificity and effect of pH and ionic strength on the kinetics of virus binding to histo-blood group antigens (HBGA)-conjugated magnetic beads. HBGAs from porcine gastric mucin (PGM) have been conjugated to magnetic beads (PGM-MB) for concentration of NoV. A GII.4 virus was used for the detailed binding ...

The binding constant (K(DOC)) between humic acid and the nitrogen-containing polycyclic aromatic compound (N-PAC), benzo[h]quinoline, was measured at varying pH levels using fluorescence quenching (FQ). Because fluorescence characteristics of benzo[h]quinoline change with pH, determination required two optimum sets of excitation and emission wavelength pairs. A simple mixing model was used ...