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1.
We report the synthesis of an electronically-tuned minimally interfering photo-affinity label (MI-PAL), a compact five-carbon tag functionalized with an alkyl diazirine and alkyne handle. MI-PAL is compatible with protein photo-conjugation, click chemistry and mass spectrometry and readily installed to complex molecules for biological target identification. 相似文献
2.
Olivier Cala Florence Guillière Isabelle Krimm 《Analytical and bioanalytical chemistry》2014,406(4):943-956
Physiological processes are mainly controlled by intermolecular recognition mechanisms involving protein–protein and protein–ligand (low molecular weight molecules) interactions. One of the most important tools for probing these interactions is high-field solution nuclear magnetic resonance (NMR) through protein-observed and ligand-observed experiments, where the protein receptor or the organic compounds are selectively detected. NMR binding experiments rely on comparison of NMR parameters of the free and bound states of the molecules. Ligand-observed methods are not limited by the protein molecular size and therefore have great applicability for analysing protein–ligand interactions. The use of these NMR techniques has considerably expanded in recent years, both in chemical biology and in drug discovery. We review here three major ligand-observed NMR methods that depend on the nuclear Overhauser effect—transferred nuclear Overhauser effect spectroscopy, saturation transfer difference spectroscopy and water–ligand interactions observed via gradient spectroscopy experiments—with the aim of reporting recent developments and applications for the characterization of protein–ligand complexes, including affinity measurements and structural determination. 相似文献
3.
《Electrochemistry communications》2005,7(5):537-540
We demonstrate for the first time the utility of nucleic acid aptamers for electrochemical detection of proteins. Highly specific and sensitive label-free detection of the target protein is achieved by combining aptamer-coated magnetic beads and chronopotentiometric stripping measurements of the captured protein (in connection to the intrinsic electroactivity of the protein). Lysozyme has thus been detected selectively in a mixture containing a large excess of six proteins and amino acids (both electroactive and non-electroactive), with a detection limit of 350 fmol (7 nM). While aptamer-based electronic sensors are in their infancy, such devices offer attractive opportunities for electrochemical detection of proteins and for developing proteomic chips. 相似文献
4.
Baltzer L 《Analytical and bioanalytical chemistry》2011,400(6):1653-1664
A new concept for protein recognition and binding is highlighted. The conjugation of small organic molecules or short peptides
to polypeptides from a designed set provides binder molecules that bind proteins with high affinities, and with selectivities
that are equal to those of antibodies. The small organic molecules or peptides need to bind the protein targets but only with
modest affinities and selectivities, because conjugation to the polypeptides results in molecules with dramatically improved
binder performance. The polypeptides are selected from a set of only sixteen sequences designed to bind, in principle, any
protein. The small number of polypeptides used to prepare high-affinity binders contrasts sharply with the huge libraries
used in binder technologies based on selection or immunization. Also, unlike antibodies and engineered proteins, the polypeptides
have unordered three-dimensional structures and adapt to the proteins to which they bind. Binder molecules for the C-reactive
protein, human carbonic anhydrase II, acetylcholine esterase, thymidine kinase 1, phosphorylated proteins, the D-dimer, and
a number of antibodies are used as examples to demonstrate that affinities are achieved that are higher than those of the
small molecules or peptides by as much as four orders of magnitude. Evaluation by pull-down experiments and ELISA-based tests
in human serum show selectivities to be equal to those of antibodies. Small organic molecules and peptides are readily available
from pools of endogenous ligands, enzyme substrates, inhibitors or products, from screened small molecule libraries, from
phage display, and from mRNA display. The technology is an alternative to established binder concepts for applications in
drug development, diagnostics, medical imaging, and protein separation. 相似文献
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《Journal of membrane science》1997,125(1):93-108
Bulk mass transfer limitations can have a significant effect on the flux and selectivity during membrane ultrafiltration. Most previous studies of these phenomena have employed the simple stagnant film analysis, but this model is unable to account for the effects of solute–solute interactions on mass transport. We have developed a generalized framework for multicomponent mass transfer that includes both thermodynamic and hydrodynamic (frictional) interactions. Thermodynamic (virial) coefficients were evaluated from osmotic pressure data for albumin (BSA) and immunoglobulins (IgG), while hydrodynamic interaction parameters were determined from filtrate flux data obtained in a stirred cell using fully retentive membranes. The protein concentration profiles in the bulk solution were evaluated by numerical solution of the governing continuity equations incorporating the multicomponent diffusive flux. This model was used to analyze flux and protein transmission data obtained for the filtration of BSA and IgG mixtures through partially permeable membranes. The model accurately predicted the large reduction in flux and BSA transmission upon addition of IgG. These effects were due to the coupling between BSA and IgG mass transfer caused by protein–protein interactions. 相似文献
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In proteins, the number of interacting pairs is usually much smaller than the number of non-interacting ones. So the imbalanced data problem will arise in the field of protein–protein interactions (PPIs) prediction. In this article, we introduce two ensemble methods to solve the imbalanced data problem. These ensemble methods combine the based-cluster under-sampling technique and the fusion classifiers. And then we evaluate the ensemble methods using a dataset from Database of Interacting Proteins (DIP) with 10-fold cross validation. All the prediction models achieve area under the receiver operating characteristic curve (AUC) value about 95%. Our results show that the ensemble classifiers are quite effective in predicting PPIs; we also gain some valuable conclusions on the performance of ensemble methods for PPIs in imbalanced data. The prediction software and all dataset employed in the work can be obtained for free at http://cic.scu.edu.cn/bioinformatics/Ensemble_PPIs/index.html. 相似文献
10.
Friedhoff P 《Analytical and bioanalytical chemistry》2005,381(1):78-80
11.
Bowater Richard P. Cobb Andrew M. Pivonkova Hana Havran Ludek Fojta Miroslav 《Monatshefte für Chemie / Chemical Monthly》2015,146(5):723-739
Monatshefte für Chemie - Chemical Monthly - This review is devoted to biophysical and electrochemical methods used for studying protein–nucleic acid (NA) interactions. The importance of... 相似文献
12.
The study of metal–protein interactions is an expanding field of research investigated by bioinorganic chemists as it has wide applications in biological systems. Very recently, it has been reported that it is possible to study metal–protein interactions by immobilizing biomolecules on metal surfaces and applying experimental approaches based on plasmonics which have usually been used to investigate protein–protein interactions. This is possible because the electronic structure of metals generates plasmons whose properties can be exploited to obtain information from biomolecules that interact not only with other molecules but also with ions in solution. One major challenge of such approaches is to immobilize the protein to be studied on a metal surface with preserved native structure. This review reports and discusses all the works that deal with such an expanding new field of application of plasmonics with specific attention to surface plasmon resonance, highlighting the advantages and drawbacks of such approaches in comparison with other experimental techniques traditionally used to study metal–protein interactions. Figure
Plasmonics is a powerful tool for the study of metal ion-protein interactions 相似文献
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14.
Studying protein–protein affinity and immobilized ligand–protein affinity interactions using MS-based methods 总被引:1,自引:0,他引:1
This review discusses the most important current methods employing mass spectrometry (MS) analysis for the study of protein
affinity interactions. The methods are discussed in depth with particular reference to MS-based approaches for analyzing protein–protein
and protein–immobilized ligand interactions, analyzed either directly or indirectly. First, we introduce MS methods for the
study of intact protein complexes in the gas phase. Next, pull-down methods for affinity-based analysis of protein–protein
and protein–immobilized ligand interactions are discussed. Presently, this field of research is often called interactomics
or interaction proteomics. A slightly different approach that will be discussed, chemical proteomics, allows one to analyze
selectivity profiles of ligands for multiple drug targets and off-targets. Additionally, of particular interest is the use
of surface plasmon resonance technologies coupled with MS for the study of protein interactions. The review addresses the
principle of each of the methods with a focus on recent developments and the applicability to lead compound generation in
drug discovery as well as the elucidation of protein interactions involved in cellular processes. The review focuses on the
analysis of bioaffinity interactions of proteins with other proteins and with ligands, where the proteins are considered as
the bioactives analyzed by MS. 相似文献
15.
Becker JS Boulyga SF Pickhardt C Becker J Buddrus S Przybylski M 《Analytical and bioanalytical chemistry》2003,375(4):561-566
Inductively coupled plasma mass spectrometry (ICP-MS) is used for phosphorus determination in protein samples. A small amount of solid protein sample (down to 1 micro g) or digest (1-10 micro L) protein solution was denatured in nitric acid and hydrogen peroxide by closed-microvessel microwave digestion. Phosphorus determination was performed with an optimized analytical method using a double-focusing sector field inductively coupled plasma mass spectrometer (ICP-SFMS) and quadrupole-based ICP-MS (ICP-QMS). For quality control of phosphorus determination a certified reference material (CRM), single cell proteins (BCR 273) with a high phosphorus content of 26.8+/-0.4 mg g(-1), was analyzed. For studies on phosphorus determination in proteins while reducing the sample amount as low as possible the homogeneity of CRM BCR 273 was investigated. Relative standard deviation and measurement accuracy in ICP-QMS was within 2%, 3.5%, 11% and 12% when using CRM BCR 273 sample weights of 40 mg, 5 mg, 1 mg and 0.3 mg, respectively. The lowest possible sample weight for an accurate phosphorus analysis in protein samples by ICP-MS is discussed. The analytical method developed was applied for the analysis of homogeneous protein samples in very low amounts [1-100 micro g of solid protein sample, e.g. beta-casein or down to 1 micro L of protein or digest in solution (e.g., tau protein)]. A further reduction of the diluted protein solution volume was achieved by the application of flow injection in ICP-SFMS, which is discussed with reference to real protein digests after protein separation using 2D gel electrophoresis.The detection limits for phosphorus in biological samples were determined by ICP-SFMS down to the ng g(-1) level. The present work discusses the figure of merit for the determination of phosphorus in a small amount of protein sample with ICP-SFMS in comparison to ICP-QMS. 相似文献
16.
New fast methods for the determination of pharmacokinetic behaviour of potential drug candidates are receiving increasing
interest. We present a new homogeneous method for the determination of drug binding and drug competition for human serum albumin
and α1-acid glycoprotein that is amenable to high-throughput-screening. It is based on selective fluorescent probes and the measurement
of fluorescence polarization. This leads to decreased interference with fluorescent drugs as compared with previously published
methods based on similar probes and the measurement of fluorescence intensity. The binding of highly fluorescent drugs that
still interfere with the probes can be measured by simply titrating the drugs in a two-component system with the serum protein.
The assay may also be used to discover strongly binding protein ligands that are interesting for drug-targeting strategies.
Additionally, binding data could be obtained from larger libraries of compounds for in silico predictive pharmacokinetics.
Figure Fluorescence polarization displacement titration of dansylsarcosine (3D-structure as insert) bound to human serum albumin
(HSA) by naproxene
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
17.
A simple electrochemical method for the determination of association constants between carbohydrates and carbohydrate-binding proteins using cyclic voltammetry (CV) is described. The binding of concanavalin A (Con A) and cholera toxin (CT) to their specific α-mannose and β-galactose derivatives self-assembled on gold electrodes is electrochemically monitored with a redox probe of K3Fe(CN)6/K4Fe(CN)6. Upon binding of the proteins to the carbohydrate-modified electrodes, the redox current in CV decreases. The binding-induced change in electrochemical signal is thus used to construct Langmuir adsorption isotherm for the carbohydrate–protein interactions and to obtain the association constants. The association constants of carbohydrate–protein interactions determined by CV ((5.8 ± 1.2) × 107 M− 1 for mannose–Con A, (2.6 ± 0.5) × 108 M− 1 for galactose-CT) were in good agreement with those measured with electrochemical impedance spectroscopy and quartz crystal microbalance. 相似文献
18.
Journal of Computer-Aided Molecular Design - Protein–protein interactions (PPIs) play crucial roles in many cellular processes and their deregulation often leads to cellular dysfunctions. One... 相似文献
19.
Andrea Sinz 《Analytical and bioanalytical chemistry》2010,397(8):3433-3440
The identification of protein–protein interactions within their physiological environment is the key to understanding biological processes at the molecular level. However, the artificial nature of in vitro experiments, with their lack of other cellular components, may obstruct observations of specific cellular processes. In vivo analyses can provide information on the processes within a cell that might not be observed in vitro. Chemical crosslinking combined with mass spectrometric analysis of the covalently connected binding partners allows us to identify interacting proteins and to map their interface regions directly in the cell. In this paper, different in vivo crosslinking strategies for deriving information on protein–protein interactions in their physiological environment are described. 相似文献