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Pharmacokinetics, brain distribution and plasma protein binding of carbamazepine and nine derivatives: new set of data for predictive in silico ADME models

Fortuna, Ana; Alves, Gilberto; Soares-da-Silva, Patrício; Falcão, Amílcar
Fonte: Elsevier Publicador: Elsevier
Tipo: Artigo de Revista Científica
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In silico approaches to predict absorption, distribution, metabolism and excretion (ADME) of new drug candidates are gaining a relevant importance in drug discovery programmes. When considering particularly the pharmacokinetics during the development of oral antiepileptic drugs (AEDs), one of the most prominent goals is designing compounds with good bioavailability and brain penetration. Thus, it is expected that in silico models able to predict these features may be applied during the early stages of AEDs discovery. The present investigation was mainly carried out in order to generate in vivo pharmacokinetic data that can be utilized for development and validation of in silico models. For this purpose, a single dose of each compound (1.4 mmol/kg) was orally administered to male CD-1 mice. After quantifying the parent compound and main metabolites in plasma and brain up to 12 h post-dosing, a non-compartmental pharmacokinetic analysis was performed and the corresponding brain/plasma ratios were calculated. Moreover the plasma protein binding was estimated in vitro applying the ultrafiltration procedure. The present in vivo pharmacokinetic characterization of the test compounds and corresponding metabolites demonstrated that the metabolism extensively compromised the in vivo activity of CBZ derivatives and their toxicity. Furthermore...

Influence of Plasma Protein Binding on Pharmacodynamics: Estimation of In Vivo Receptor Affinities of beta Blockers Using a New Mechanism-Based PK-PD Modelling Approach

STEEG, T. J. Van; BORALLI, V. B.; KREKELS, E. H. J.; SLIJKERMAN, P.; FREIJER, J.; DANHOF, M.; LANGE, E. C. M. De
Fonte: JOHN WILEY & SONS INC Publicador: JOHN WILEY & SONS INC
Tipo: Artigo de Revista Científica
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The objective of this investigation was to examine in a systematic manner the influence of plasma protein binding on in vivo pharmacodynamics. Comparative pharmacokinetic-pharmacodynamic studies with four beta blockers were performed in conscious rats, using heart rate under isoprenaline-induced tachycardia as a pharmacodynamic endpoint. A recently proposed mechanism-based agonist-antagonist interaction model was used to obtain in vivo estimates of receptor affinities (K(B),(vivo)). These values were compared with in vitro affinities (K(B),(vitro)) on the basis of both total and free drug concentrations. For the total drug concentrations, the K(B),(vivo) estimates were 26, 13, 6.5 and 0.89 nM for S(-)-atenolol, S(-)-propranolol, S(-)-metoprolol and timolol. The K(B),(vivo) estimates on the basis of the free concentrations were 25, 2.0, 5.2 and 0.56 nM, respectively. The K(B),(vivo)-K(B),(vitro) correlation for total drug concentrations clearly deviated from the line of identity, especially for the most highly bound drug S(-)-propranolol (ratio K(B),(vivo)/K(B),(vitro) similar to 6.8). For the free drug, the correlation approximated the line of identity. Using this model, for beta-blockers the free plasma concentration appears to be the best predictor of in vivo pharmacodynamics. (C) 2008 Wiley-Liss...

Plasma levels of transthyretin and retinol-binding protein in child-a cirrhotic patients in relation to protein-calorie status and plasma amino acids, zinc, vitamin a and plasma thyroid hormones

Calamita, Zamir; Dichi, Isaías; Papini-Berto, Sílvia J.; Dichi, Jane B.; Angeleli, Aparecida Y.O.; Vannucchi, Hélio; Caramori, Carlos; Burini, Roberto Carlos
Tipo: Artigo de Revista Científica Formato: 139-147
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Transthyretin and retinal-binding protein are sensitive markers of acute protein-calorie malnutrition both for early diagnosis and dietary evaluation. A preliminary study showed that retinal-binding protein is the most sensitive marker of protein-calorie malnutrition in cirrhotic patients, even those with the mild form of the disease (Child A). However, in addition to being affected by protein-calorie malnutrition, the levels of these short half-life-liver-produced proteins are also influenced by other factors of a nutritional (zinc, tryptophan, vitamin A, etc) and non-nutritional (sex, aging, hormones, renal and liver functions and inflammatory activity) nature. These interactions were investigated in 11 adult male patients (49.9 ± 9.2 years of age) with alcoholic cirrhosis (Child-Pugh grade A) and with normal renal function. Both transthyretin and retinol binding protein were reduced below normal levels in 55% of the patients, in close agreement with their plasma levels of retinal. In 67% of the patients (4/6), the reduced levels of transthyretin and retinal-binding protein were caused by altered liver function and in 50% (3/6) they were caused by protein-calorie malnutrition. Thus, the present data, taken as a whole, indicate that reduced transthyretin and retinal-binding protein levels in mild cirrhosis of the liver are mainly due to liver failure and/or vitamin A status rather than representing an isolated protein-calorie malnutrition indicator.

Mapping eIF5A binding sites for Dys1 and Lia1: In vivo evidence for regulation of eIF5A hypusination

Thompson, Gloria M.; Cano, Veridiana S.P.; Valentini, Sandro R.
Tipo: Artigo de Revista Científica Formato: 464-468
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The evolutionarily conserved factor eIF5A is the only protein known to undergo hypusination, a unique posttranslational modification triggered by deoxyhypusine synthase (Dys1). Although eIF5A is essential for cell viability, the function of this putative translation initiation factor is still obscure. To identify eIF5A-binding proteins that could clarify its function, we screened a two-hybrid library and identified two eIF-5A partners in S. cerevisiae: Dys1 and the protein encoded by the gene YJR070C, named Lia1 (Ligand of eIF5A). The interactions were confirmed by GST pulldown. Mapping binding sites for these proteins revealed that both eIF5A domains can bind to Dys1, whereas the C-terminal domain is sufficient to bind Lia1. We demonstrate for the first time in vivo that the N-terminal ?-helix of Dys1 can modulate enzyme activity by inhibiting eIF5A interaction. We suggest that this inhibition be abrogated in the cell when hypusinated and functional eIF5A is required. © 2003 Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.

Interaction of Bacillus thuringiensis Cry1 and Vip3A proteins with Spodoptera frugiperda midgut binding sites

Sena, Janete A. D.; Hernández-Rodríguez, Carmen Sara; Ferré, Juan
Tipo: Artigo de Revista Científica Formato: 2236-2237
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Vip3Aa, Vip3Af, Cry1Ab, and Cry1Fa were tested for their toxicities and binding interactions. Vip3A proteins were more toxic than Cry1 proteins. Binding assays showed independent specific binding sites for Cry1 and Vip3A proteins. Cry1Ab and Cry1Fa competed for the same binding sites, whereas Vip3Aa competed for those of Vip3Af. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Diclofenac plasma protein binding: PK-PD modelling in cardiac patients submitted to cardiopulmonary bypass

Fonte: Associação Brasileira de Divulgação Científica Publicador: Associação Brasileira de Divulgação Científica
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/03/1997 Português
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Twenty-four surgical patients of both sexes without cardiac, hepatic, renal or endocrine dysfunctions were divided into two groups: 10 cardiac surgical patients submitted to myocardial revascularization and cardiopulmonary bypass (CPB), 3 females and 7 males aged 65 ± 11 years, 74 ± 16 kg body weight, 166 ± 9 cm height and 1.80 ± 0.21 m2 body surface area (BSA), and control, 14 surgical patients not submitted to CPB, 11 female and 3 males aged 41 ± 14 years, 66 ± 14 kg body weight, 159 ± 9 cm height and 1.65 ± 0.16 m2 BSA (mean ± SD). Sodium diclofenac (1 mg/kg, im Voltaren 75® twice a day) was administered to patients in the Recovery Unit 48 h after surgery. Venous blood samples were collected during a period of 0-12 h and analgesia was measured by the visual analogue scale (VAS) during the same period. Plasma diclofenac levels were measured by high performance liquid chromatography. A two-compartment open model was applied to obtain the plasma decay curve and to estimate kinetic parameters. Plasma diclofenac protein binding decreased whereas free plasma diclofenac levels were increased five-fold in CPB patients. Data obtained for analgesia reported as the maximum effect (EMAX) were: 25% VAS (CPB) vs 10% VAS (control), P<0.05...

In vitro protein binding of cefonicid and cefuroxime in adult and neonatal sera.

Benson, J M; Boudinot, F D; Pennell, A T; Cunningham, F E; DiPiro, J T
Tipo: text
Publicado em /06/1993 Português
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The levels of in vitro protein binding of cefonicid and cefuroxime in human adult and neonatal sera were compared. Binding parameters for each drug were determined within the concentration range of 25 to 3,000 micrograms/ml. Cefonicid exhibited concentration-dependent protein binding in both types of sera, with more extensive binding in adult serum at all concentrations. Two classes of binding sites were found: a high-affinity, saturable site and a low-affinity, nonspecific site. Cefuroxime also showed two-class, concentration-dependent protein binding in adult serum, but binding in neonatal serum was to a single class and was independent of drug concentration. Parameters for class 1 binding sites for cefonicid indicated one binding site per albumin molecule in both adult and neonatal sera, with association constants of 7.0 x 10(4) and 1.3 x 10(4) M-1, respectively. These parameters were also derived for cefuroxime in adult serum and were 0.15 and 7.1 x 10(4) M-1, respectively. In neonatal serum, the combined value (number of binding sites per molecule x equilibrium association constant) was similar to combined values calculated for class 2 binding sites for cefuroxime in adult serum and for cefonicid in neonatal serum (287 versus 195 and 261 M-1...

Cellular or viral protein binding to a cytomegalovirus promoter transcription initiation site: effects on transcription.

Macias, M P; Huang, L; Lashmit, P E; Stinski, M F
Tipo: text
Publicado em /06/1996 Português
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We have previously shown that the IE2 protein of human cytomegalovirus (CMV) represses its own synthesis by binding to the major immediate-early promoter (M. P. Macias and M. F. Stinski, Proc. Natl. Acad. Sci. USA 90:707-711, 1993). The binding of a viral protein (IE2) and a cellular protein in the region of the transcription start site was investigated by site-specific mutational analysis and electrophoretic mobility shift assay. The viral protein and the cellular protein require different but adjacent core DNA sequence elements for binding. In situ chemical footprinting analysis of DNA-protein interactions with purified CMV IE2 protein or HeLa cell nuclear extracts demonstrated binding sites that overlap the transcription start site. The IE2 protein footprint was between bp -15 and +2, relative to the transcription start site, and the cellular protein was between bp -16 and +7. The ability of the unknown human cellular protein of approximately 150 kDa to bind the CMV major immediate-early promoter correlates with an increase in the level of transcription efficiency. Mutations in the core DNA sequence element for cellular protein binding significantly reduced the level of in vitro transcription efficiency. Mutations upstream and downstream of the core sequence moderately reduced the transcription efficiency level. Negative autoregulation of the CMV promoter by the viral IE2 protein may involve both binding to the DNA template and interference with the function of a cellular protein that binds to the transcription start site and enhances transcription efficiency.

Protein Binding by Specific Receptors on Human Placenta, Murine Placenta, and Suckling Murine Intestine in Relation to Protein Transport across These Tissues

Gitlin, Jonathan D.; Gitlin, David
Tipo: text
Publicado em /11/1974 Português
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Human, rat, and mouse placentas and rat and mouse intestines were homogenized in buffered saline, and fraction consisting primarily of cell membranes was separated from each of the homogenates by differential centrifugation. Human, bovine, and guinea pig IgG, and human IgE, Bence-Jones protein, serum albumin, insulin, and growth hormone were labeled with 131I or 125I, and the binding of these proteins by the cell membrane fractions was investigated. Rat and mouse sucklings were given labeled proteins intragastrically, and the amount of each protein absorbed after a given interval of time was determined. It was found that the degree and specificity of protein binding by the cell membrane fractions from human and murine placentas strikingly paralleled the relative rate and specificity of protein transport from mother to fetus in the respective species at or near term. Similarly, the degree and specificity of protein binding by the cell membrane fractions from suckling rat and mouse intestines tended to parallel the rate and specificity of protein absorption from the gastrointestinal tract in these animals. However, some discordance between protein binding and protein transport was also observed. The data suggest that: (a) the binding of a protein by specific receptors on cell membranes may be a necessary first step in the transcellular transport of the protein; (b) specific protein binding by cell receptors does not ensure the transport of that protein across the tissue barrier; and (c) specific transport mechanisms other than or in addition to specific cell membrane receptors are involved in the active transport of proteins across the human or murine placenta or the suckling murine intestine.

Prediction of interacting single-stranded RNA bases by protein binding patterns

Shulman-Peleg, Alexandra; Shatsky, Maxim; Nussinov, Ruth; Wolfson, Haim J.
Tipo: text
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Prediction of protein-RNA interactions at the atomic level of detail is crucial for our ability to understand and interfere with processes such as gene expression and regulation. Here, we investigate protein binding pockets that accommodate extruded nucleotides not involved in RNA base pairing. We observed that most of the protein interacting nucleotides are part of a consecutive fragment of at least two nucleotides, whose rings have significant interactions with the protein. Many of these share the same protein binding cavity and more than 30% of such pairs are ?-stacked. Since these local geometries can not be inferred from the nucleotide identities, we present a novel framework for their prediction from the properties of protein binding sites. First, we present a classification of known RNA nucleotide and dinucleotide protein binding sites and identify the common types of shared 3D physico-chemical binding patterns. These are recognized by a new classification methodology which is based on spatial multiple alignment. The shared patterns reveal novel similarities between dinucleotide binding sites of proteins with different overall sequences, folds and functions. Given a protein structure, we use these patterns for the prediction of its RNA dinucleotides binding sites. Based on the binding modes of these nucleotides...

Peptide and Protein Binding in the Axial Channel of Hsp104: INSIGHTS INTO THE MECHANISM OF PROTEIN UNFOLDING*

Lum, Ronnie; Niggemann, Monika; Glover, John R.
Fonte: American Society for Biochemistry and Molecular Biology Publicador: American Society for Biochemistry and Molecular Biology
Tipo: text
Publicado em 31/10/2008 Português
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The AAA+ molecular chaperone Hsp104 mediates the extraction of proteins from aggregates by unfolding and threading them through its axial channel in an ATP-driven process. An Hsp104-binding peptide selected from solid phase arrays enhanced the refolding of a firefly luciferase-peptide fusion protein. Analysis of peptide binding using tryptophan fluorescence revealed two distinct binding sites, one in each AAA+ module of Hsp104. As a further indication of the relevance of peptide binding to the Hsp104 mechanism, we found that it competes with the binding of a model unfolded protein, reduced carboxymethylated ?-lactalbumin. Inactivation of the pore loops in either AAA+ module prevented stable peptide and protein binding. However, when the loop in the first AAA+ was inactivated, stimulation of ATPase turnover in the second AAA+ module of this mutant was abolished. Drawing on these data, we propose a detailed mechanistic model of protein unfolding by Hsp104 in which an initial unstable interaction involving the loop in the first AAA+ module simultaneously promotes penetration of the substrate into the second axial channel binding site and activates ATP turnover in the second AAA+ module.

Structure-based Protocol for Identifying Mutations that Enhance Protein-Protein Binding Affinities

Sammond, Deanne W.; Eletr, Ziad M.; Purbeck, Carrie; Kimple, Randall J.; Siderovski, David P.; Kuhlman, Brian
Tipo: text
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The ability to manipulate protein binding affinities is important for the development of proteins as biosensors, industrial reagents, and therapeutics. We have developed a structure-based method to rationally predict single mutations at protein-protein interfaces that enhance binding affinities. The protocol is based on the premise that increasing buried hydrophobic surface area and/or reducing buried hydrophilic surface area will generally lead to enhanced affinity if large steric clashes are not introduced and buried polar groups are not left without a hydrogen bond partner. The procedure selects affinity enhancing point mutations at the protein-protein interface using three criteria: 1) the mutation must be from a polar amino acid to a non-polar amino acid or from a non-polar amino acid to a larger non-polar amino acid, 2) the free energy of binding as calculated with the Rosetta protein modeling program should be more favorable than the free energy of binding calculated for the wild type complex and 3) the mutation should not be predicted to significantly destabilize the monomers. The Rosetta energy function emphasizes short-range interactions: steric repulsion, Van der Waals forces, hydrogen bonding, and an implicit solvation model that penalizes placing atoms adjacent to polar groups. The performance of the computational protocol was experimentally tested on two separate protein complexes; G?i1 from the heterotrimeric G-protein system bound to the RGS14 GoLoco motif...

RsiteDB: a database of protein binding pockets that interact with RNA nucleotide bases

Shulman-Peleg, Alexandra; Nussinov, Ruth; Wolfson, Haim J.
Fonte: Oxford University Press Publicador: Oxford University Press
Tipo: text
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We present a new database and an on-line search engine, which store and query the protein binding pockets that interact with single-stranded RNA nucleotide bases. The database consists of a classification of binding sites derived from protein–RNA complexes. Each binding site is assigned to a cluster of similar binding sites in other protein–RNA complexes. Cluster members share similar spatial arrangements of physico–chemical properties, thus can reveal novel similarity between proteins and RNAs with different sequences and folds. The clusters provide 3D consensus binding patterns important for protein–nucleotide recognition. The database search engine allows two types of useful queries: first, given a PDB code of a protein–RNA complex, RsiteDB can detail and classify the properties of the protein binding pockets accommodating extruded RNA nucleotides not involved in local RNA base pairing. Second, given an unbound protein structure, RsiteDB can perform an on-line structural search against the constructed database of 3D consensus binding patterns. Regions similar to known patterns are predicted to serve as binding sites. Alignment of the query to these patterns with their corresponding RNA nucleotides allows making unique predictions of the protein–RNA interactions at the atomic level of detail. This database is accessable at http://bioinfo3d.cs.tau.ac.il/RsiteDB.

Tethered-Hopping Model for Protein-DNA Binding and Unbinding Based on Sox2-Oct1-Hoxb1 Ternary Complex Simulations

Lian, Peng; Angela Liu, Limin; Shi, Yongxiang; Bu, Yuxiang; Wei, Dongqing
Fonte: The Biophysical Society Publicador: The Biophysical Society
Tipo: text
Publicado em 07/04/2010 Português
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The sliding and hopping models encapsulate the essential protein-DNA binding process for binary complex formation and dissociation. However, the effects of a cofactor protein on the protein-DNA binding process that leads to the formation of a ternary complex remain largely unknown. Here we investigate the effect of the cofactor Sox2 on the binding and unbinding of Oct1 with the Hoxb1 control element. We simulate the association of Oct1 with Sox2-Hoxb1 using molecular dynamics simulations, and the dissociation of Oct1 from Sox2-Hoxb1 using steered molecular dynamics simulations, in analogy to a hopping event of Oct1. We compare the kinetic and thermodynamic properties of three model complexes (the wild-type and two mutants) in which the Oct1-DNA base-specific interactions or the Sox2-Oct1 protein-protein interactions are largely abolished. We find that Oct1-DNA base-specific interactions contribute significantly to the total interaction energy of the ternary complex, and that nonspecific Oct1-DNA interactions are sufficient for driving the formation of the protein-DNA interface. The Sox2-Oct1 protein-protein binding interface is largely hydrophobic, with remarkable shape complementarity. This interface promotes the formation of the ternary complex and slows the dissociation of Oct1 from its DNA-binding site. We propose a simple two-step reaction model of protein-DNA binding...

Improved Ligand-Protein Binding Affinity Predictions Using Multiple Binding Modes

Stjernschantz, Eva; Oostenbrink, Chris
Fonte: The Biophysical Society Publicador: The Biophysical Society
Tipo: text
Publicado em 02/06/2010 Português
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Accurate ligand-protein binding affinity prediction, for a set of similar binders, is a major challenge in the lead optimization stage in drug development. In general, docking and scoring functions perform unsatisfactorily in this application. Docking calculations, followed by molecular dynamics simulations and free energy calculations can be applied to improve the predictions. However, for targets with large, flexible binding sites, with no experimentally determined binding modes for a set of ligands, insufficient sampling can decrease the accuracy of the free energy calculations. Cytochrome P450s, a protein family of major importance for drug metabolism, is an example of a challenging target for binding affinity predictions. As a result, the choice of starting structure from the docking solutions becomes crucial. In this study, an iterative scheme is introduced that includes multiple independent molecular dynamics simulations to obtain weighted ensemble averages to be used in the linear interaction energy method. The proposed scheme makes the initial pose selection less crucial for further simulation, as it automatically calculates the relative weights of the various poses. It also properly takes into account the possibility that multiple binding modes contribute similarly to the overall affinity...

Protein Binding: Do We Ever Learn??

Zeitlinger, Markus A.; Derendorf, Hartmut; Mouton, Johan W.; Cars, Otto; Craig, William A.; Andes, David; Theuretzbacher, Ursula
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: text
Publicado em /07/2011 Português
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Although the influence of protein binding (PB) on antibacterial activity has been reported for many antibiotics and over many years, there is currently no standardization for pharmacodynamic models that account for the impact of protein binding of antimicrobial agents in vitro. This might explain the somewhat contradictory results obtained from different studies. Simple in vitro models which compare the MIC obtained in protein-free standard medium versus a protein-rich medium are prone to methodological pitfalls and may lead to flawed conclusions. Within in vitro test systems, a range of test conditions, including source of protein, concentration of the tested antibiotic, temperature, pH, electrolytes, and supplements may influence the impact of protein binding. As new antibiotics with a high degree of protein binding are in clinical development, attention and action directed toward the optimization and standardization of testing the impact of protein binding on the activity of antibiotics in vitro become even more urgent. In addition, the quantitative relationship between the effects of protein binding in vitro and in vivo needs to be established, since the physiological conditions differ. General recommendations for testing the impact of protein binding in vitro are suggested.

Refining Vancomycin Protein Binding Estimates: Identification of Clinical Factors That Influence Protein Binding?

Butterfield, Jill M.; Patel, Nimish; Pai, Manjunath P.; Rosano, Thomas G.; Drusano, George L.; Lodise, Thomas P.
Fonte: American Society for Microbiology Publicador: American Society for Microbiology
Tipo: text
Publicado em /09/2011 Português
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While current data indicate only free (unbound) drug is pharmacologically active and is most predictive of response, pharmacodynamic studies of vancomycin have been limited to measurement of total concentrations. The protein binding of vancomycin is thought to be approximately 50%, but considerable variability surrounds this estimate. The present study sought to determine the extent of vancomycin protein binding, to identify factors that modulate its binding, and to create and validate a prediction tool to estimate the extent of protein binding based on individual clinical factors. This single-site prospective cohort study included hospitalized adult patients treated with vancomycin and with a vancomycin serum concentration determination available. Linear regression was used to predict the free vancomycin concentration (f[vanco]) and to determine the clinical factors modulating vancomycin protein binding. Among the 50 patients in the study, the mean protein binding was 41.5%. The strongest predictor of f[vanco] was the total vancomycin concentration (total [vanco]), and this was modified by dialysis and total protein of ?6.7 g/dl as covariates. The algebraic expression from the final prediction model was f[vanco] = 0.643 + 0.560 × total [vanco] ? {0.067 × total [vanco] × D} ? {0.071 × total [vanco] × TP} where D = 1 if dialysis dependent or 0 if not dialysis dependent...

PBSword: a web server for searching similar protein–protein binding sites

Pang, Bin; Kuang, Xingyan; Zhao, Nan; Korkin, Dmitry; Shyu, Chi-Ren
Fonte: Oxford University Press Publicador: Oxford University Press
Tipo: text
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PBSword is a web server designed for efficient and accurate comparisons and searches of geometrically similar protein–protein binding sites from a large-scale database. The basic idea of PBSword is that each protein binding site is first represented by a high-dimensional vector of ‘visual words’, which characterizes both the global and local shape features of the binding site. It then uses a scalable indexing technique to search for those binding sites whose visual words representations are similar to that of the query binding site. Our system is able to return ranked results of binding sites in short time from a database of 194?322 domain–domain binding sites. PBSword supports query by protein ID and by new structures uploaded by users. PBSword is a useful tool to investigate functional connections among proteins based on the local structures of binding site and has potential applications to protein–protein docking and drug discovery. The system is hosted at http://pbs.rnet.missouri.edu.

Regulation of protein-protein binding by coupling between phosphorylation and intrinsic disorder: analysis of human protein complexes

Nishi, Hafumi; Fong, Jessica; Chang, Christiana; Teichmann, Sarah A.; Panchenko, Anna R.
Tipo: text
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Phosphorylation offers a dynamic way to regulate protein activity, subcellular localization, and stability. The majority of signaling pathways involve an extensive set of protein-protein interactions, and phosphorylation is widely used to regulate protein-protein binding by affecting the stability, kinetics and specificity of interactions. Previously it was found that phosphorylation sites tend to be located on protein-protein binding interfaces and may orthosterically modulate the strength of interactions. Here we studied the effect of phosphorylation on protein binding in relation to intrinsic disorder for different types of human protein complexes with known structure of binding interface. Our results suggest that the processes of phosphorylation, binding and disorder-order transitions are coupled to each other, with about one quarter of all disordered interface Ser/Thr/Tyr sites being phosphorylated. Namely, residue site disorder and interfacial states significantly affect the phosphorylation of serine and to a lesser extent of threonine. Tyrosine phosphorylation might not be directly associated with binding through disorder, and is often observed in ordered interface regions which are not predicted to be disordered in the unbound state. We analyze possible mechanisms of how phosphorylation might regulate protein-protein binding via intrinsic disorder...

Modeling Spatial Correlation of DNA Deformation: DNA Allostery in Protein Binding

Xu, Xinliang; Ge, Hao; Gu, Chan; Gao, Yi Qin; Wang, Siyuan S.; Thio, Beng Joo Reginald; Hynes, James T.; Xie, Xiaoliang Sunney; Cao, Jianshu
Fonte: American Chemical Society Publicador: American Chemical Society
Tipo: Artigo de Revista Científica
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We report a study of DNA deformations using a coarse-grained mechanical model and quantitatively interpret the allosteric effects in protein–DNA binding affinity. A recent single-molecule study (Kim et al. Science 2013, 339, 816) showed that when a DNA molecule is deformed by specific binding of a protein, the binding affinity of a second protein separated from the first protein is altered. Experimental observations together with molecular dynamics simulations suggested that the origin of the DNA allostery is related to the observed deformation of DNA’s structure, in particular, the major groove width. To unveil and quantify the underlying mechanism for the observed major groove deformation behavior related to the DNA allostery, here we provide a simple but effective analytical model where DNA deformations upon protein binding are analyzed and spatial correlations of local deformations along the DNA are examined. The deformation of the DNA base orientations, which directly affect the major groove width, is found in both an analytical derivation and coarse-grained Monte Carlo simulations. This deformation oscillates with a period of 10 base pairs with an amplitude decaying exponentially from the binding site with a decay length (l_D approx10) base pairs as a result of the balance between two competing terms in DNA base-stacking energy. This length scale is in agreement with that reported from the single-molecule experiment. Our model can be reduced to the worm-like chain form at length scales larger than (l_P) but is able to explain DNA’s mechanical properties on shorter length scales...