Functional specialization of CK2 isoforms and characterization of isoform-specific binding partners

2001 ◽  
pp. 21-29
Author(s):  
David W. Litchfield ◽  
Denis G. Bosc ◽  
David A. Canton ◽  
Ronald B. Saulnier ◽  
Greg Vilk ◽  
...  
Keyword(s):  
Specific Binding ◽  
Functional Specialization ◽  
Binding Partners

scholarly journals Relative interfacial cleavage energetics of protein complexes revealed by surface collisions

2019 ◽  
Vol 116 (17) ◽  
pp. 8143-8148 ◽  
Author(s):  
Sophie R. Harvey ◽  
Justin T. Seffernick ◽  
Royston S. Quintyn ◽  
Yang Song ◽  
Yue Ju ◽  
...  
Keyword(s):  
Computational Models ◽  
Protein Complexes ◽  
Specific Binding ◽  
Native Mass Spectrometry ◽  
Binding Affinities ◽  
Training Set ◽  
Subunit Interactions ◽  
Binding Partners ◽  
Relative Binding

To fulfill their biological functions, proteins must interact with their specific binding partners and often function as large assemblies composed of multiple proteins or proteins plus other biomolecules. Structural characterization of these complexes, including identification of all binding partners, their relative binding affinities, and complex topology, is integral for understanding function. Understanding how proteins assemble and how subunits in a complex interact is a cornerstone of structural biology. Here we report a native mass spectrometry (MS)-based method to characterize subunit interactions in globular protein complexes. We demonstrate that dissociation of protein complexes by surface collisions, at the lower end of the typical surface-induced dissociation (SID) collision energy range, consistently cleaves the weakest protein:protein interfaces, producing products that are reflective of the known structure. We present here combined results for multiple complexes as a training set, two validation cases, and four computational models. We show that SID appearance energies can be predicted from structures via a computationally derived expression containing three terms (number of residues in a given interface, unsatisfied hydrogen bonds, and a rigidity factor).

scholarly journals Ultrafiltration Method for Plasma Protein Binding Studies and Its Limitations

Processes ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 382
Author(s):  
Camelia-Maria Toma ◽  
Silvia Imre ◽  
Camil-Eugen Vari ◽  
Daniela-Lucia Muntean ◽  
Amelia Tero-Vescan
Keyword(s):  
Protein Binding ◽  
Plasma Protein Binding ◽  
Plasma Protein ◽  
Specific Binding ◽  
Critical Role ◽  
Volume Ratio ◽  
Binding Studies ◽  
Experimental Conditions ◽  
Key Aspects

Plasma protein binding plays a critical role in drug therapy, being a key part in the characterization of any compound. Among other methods, this process is largely studied by ultrafiltration based on its advantages. However, the method also has some limitations that could negatively influence the experimental results. The aim of this study was to underline key aspects regarding the limitations of the ultrafiltration method, and the potential ways to overcome them. The main limitations are given by the non-specific binding of the substances, the effect of the volume ratio obtained, and the need of a rigorous control of the experimental conditions, especially pH and temperature. This review presents a variety of methods that can hypothetically reduce the limitations, and concludes that ultrafiltration remains a reliable method for the study of protein binding. However, the methodology of the study should be carefully chosen.

scholarly journals Identification of P-glycoprotein co-fractionating proteins and specific binding partners in rat brain microvessels

2015 ◽  
Vol 134 (2) ◽  
pp. 200-210 ◽  
Author(s):  
Margaret E. Tome ◽  
Charles P. Schaefer ◽  
Leigh M. Jacobs ◽  
Yifeng Zhang ◽  
Joseph M. Herndon ◽  
...  
Keyword(s):  
Rat Brain ◽  
Specific Binding ◽  
Binding Partners ◽  
Brain Microvessels ◽  
P Glycoprotein

A characterization of heam uptake and intracellular distribution by isolated hepatocytes

1985 ◽  
Vol 5 (7) ◽  
pp. 609-614 ◽  
Author(s):  
A. Lodola
Keyword(s):  
Endoplasmic Reticulum ◽  
Specific Binding ◽  
Isolated Hepatocytes ◽  
Intracellular Distribution ◽  
Distribution Data ◽  
Temperature Dependent ◽  
Rat Hepatocyte ◽  
Isolated Rat ◽  
Endoplasmic Reticulum Fraction

The uptake and intracellular distribution of haem by isolated rat hepatocyte suspensions was studied. An increase in cell haem content occurred after a challenge with 5, 10 or 20 μM haem, supplied as methaemalbumin. The rate of haem uptake was temperature dependent; no non-specific binding occurred. Intracellular haem distribution data are consistent with a rapid association of haem with the endoplasmic reticulum fraction prior to its accumulation in the cytosol and at the mitochondrion.

scholarly journals Evaluation of Radiolabeled Girentuximab In Vitro and In Vivo

2018 ◽  
Vol 11 (4) ◽  
pp. 132 ◽  
Author(s):  
Tais Basaco ◽  
Stefanie Pektor ◽  
Josue Bermudez ◽  
Niurka Meneses ◽  
Manfred Heller ◽  
...  
Keyword(s):  
Specific Binding ◽  
Sodium Ascorbate ◽  
Tumor Uptake ◽  
Tetraacetic Acid ◽  
In Vivo Experiments ◽  
Caix Expression ◽  
Tetraazacyclododecane Tetraacetic Acid

Girentuximab (cG250) targets carbonic anhydrase IX (CAIX), a protein which is expressed on the surface of most renal cancer cells (RCCs). cG250 labeled with 177Lu has been used in clinical trials for radioimmunotherapy (RIT) of RCCs. In this work, an extensive characterization of the immunoconjugates allowed optimization of the labeling conditions with 177Lu while maintaining immunoreactivity of cG250, which was then investigated in in vitro and in vivo experiments. cG250 was conjugated with S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane tetraacetic acid (DOTA(SCN)) by using incubation times between 30 and 90 min and characterized by mass spectrometry. Immunoconjugates with five to ten DOTA(SCN) molecules per cG250 molecule were obtained. Conjugates with ratios less than six DOTA(SCN)/cG250 had higher in vitro antigen affinity, both pre- and postlabeling with 177Lu. Radiochemical stability increased, in the presence of sodium ascorbate, which prevents radiolysis. The immunoreactivity of the radiolabeled cG250 tested by specific binding to SK-RC-52 cells decreased when the DOTA content per conjugate increased. The in vivo tumor uptake was < 10% ID/g and independent of the total amount of protein in the range between 5 and 100 µg cG250 per animal. Low tumor uptake was found to be due to significant necrotic areas and heterogeneous CAIX expression. In addition, low vascularity indicated relatively poor accessibility of the CAIX target.

Characterization of a specific binding site for angiotensin II in chicken liver

1997 ◽  
Vol 75 (6) ◽  
pp. 568-575 ◽  
Author(s):  
R Bouley ◽  
M Gosselin ◽  
H Plante ◽  
G Servant ◽  
J Pérodin ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Binding Site ◽  
Specific Binding ◽  
Chicken Liver

Characterization of muscarinic cholinergic receptors in cat tracheal gland cells

1986 ◽  
Vol 61 (4) ◽  
pp. 1375-1382 ◽  
Author(s):  
D. J. Culp ◽  
M. G. Marin
Keyword(s):  
Muscarinic Receptors ◽  
Specific Binding ◽  
Isolated Cells ◽  
High Affinity ◽  
Gland Cells ◽  
Beta Adrenergic Agonists ◽  
Muscarinic Cholinergic ◽  
Dose Dependent ◽  
Nanomolar Range

Studies of airway glands indicate a muscarinic cholinergic regulation of secretion. Because of the cellular complexity of the airways, receptor characterization in whole tissue is unfeasible. Therefore, we utilized homogenates of disaggregated gland cells isolated from cat trachea and the muscarinic antagonist [1–3H]quinuclidinyl benzilate ([3H]QNB) to characterize glandular muscarinic receptors. Receptors of isolated cells were functionally intact as assessed by carbachol (10(-4) M) stimulation of O2 consumption 86 +/- 6% (+/- SE, n = 20). Stimulation was dose dependent (mean effective concentration = 3.5 microM), inhibited by atropine [dissociation constant (KD) = 4.2 nM] but not phentolamine nor propranolol. Specific binding of [3H]QNB to cell homogenates was saturable, of high affinity (KD = 36 pM) and to a single population of receptors. Maximum binding was 58 fmol/10(6) cells or about 35,000 receptors per cell. Estimated affinities for muscarinic agents were in the micromolar range for agonists and nanomolar range for antagonists. Histamine, alpha-adrenergic, and beta-adrenergic agonists and antagonists did not inhibit specific binding. These results suggest that muscarinic receptors on tracheal gland cells are of high affinity and density.

Characterization of the exposed carbohydrates on the surface membrane of adult Schistosoma mansoni by analysis of lectin binding

Parasitology ◽  
1980 ◽  
Vol 81 (1) ◽  
pp. 1-15 ◽  
Author(s):  
A. J. G. Simpson ◽  
S. R. Smithers
Keyword(s):  
Schistosoma Mansoni ◽  
Adult Male ◽  
Wheat Germ ◽  
Ricinus Communis ◽  
Specific Binding ◽  
Lectin Binding ◽  
Surface Membrane ◽  
Soybean Agglutinin ◽  
Peanut Agglutinin

SUMMARYThe surface architecture of adult male Schistosoma mansoni was explored using a range of lectins with differing carbohydrate specificities. Highest specific binding was achieved with concanavalin A and the agglutinin of molecular weight 60000 from Ricinus communis; the binding of wheat germ agglutinin was mostly non-specific. Small amounts of peanut agglutinin and soybean agglutinin binding were observed and the binding of these lectins was increased by pre-treating the parasite with neuraminidase. The fucose binding protein of Lotus tetragonolobus failed to bind. These results indicate that mannose and/or glucose, galactose, N-acetyl-glucosamine, N-acetyl-galactosamine and sialic acid are exposed on the surface of the adult male schistosome.

Characterization of specific binding of atrial natriuretic peptide (ANP) to rat PC12 phaeochromocytoma cells

Life Sciences ◽  
1988 ◽  
Vol 43 (24) ◽  
pp. 2035-2042 ◽  
Author(s):  
Aicha Boumezrag ◽  
Fiona Lyall ◽  
Julian A.T. Dow
Keyword(s):  
Atrial Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Specific Binding ◽  
Atrial Natriuretic
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