The use of a prostacyclin analogue, [3H]iloprost, for studying prostacyclin-binding sites on human platelets and neuronal hybrid cells

1984 ◽  
Vol 4 (11) ◽  
pp. 941-948 ◽  
Author(s):  
Jean M. Hall ◽  
Philip G. Strange
Keyword(s):  
Ligand Binding ◽  
Binding Sites ◽  
Human Platelets ◽  
Hybrid Cells ◽  
Binding Properties ◽  
Prostacyclin Analogue ◽  
Camp Synthesis ◽  
Stimulation Of

The stable prostacyclin analogue [3H]iloprost has been used for labelling prostacyclin-binding sites on human platelets and NCB-20 neuronal hybrid cells. The ligand-binding properties of the sites have been determined and correlate well with stimulation of cAMP synthesis in NCB-20 cells and inhibition of aggregation in human platelets.

scholarly journals Affinity for (3H)Iloprost Binding Sites and cAMP Synthesis Activity of a 3-Oxa-methano Prostaglandin I1 Analog, SM-10906, in Human Platelets and Endothelial Cells.

1997 ◽  
Vol 74 (1) ◽  
pp. 37-43 ◽  
Author(s):  
Takashi Hiroi ◽  
Keiko Maruyama ◽  
Kaoru Hattori ◽  
Toshio Ohnuki ◽  
Takafumi Nagatomo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Binding Sites ◽  
Human Platelets ◽  
Camp Synthesis ◽  
Synthesis Activity

Studies on the Binding of 3H-SR121566, an Inhibitor of Gp IIb-IIIa Activation

2001 ◽  
Vol 85 (04) ◽  
pp. 702-709 ◽  
Author(s):  
P. Savi ◽  
G. Zamboni ◽  
O. Rescanières ◽  
J. M. Herbert
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Platelet Aggregation ◽  
Binding Sites ◽  
Human Platelets ◽  
Gamma Chain ◽  
Activated Platelets ◽  
Fibrinogen Binding ◽  
Direct Competition ◽  
Stimulation Of

SummarySR121566 is a new synthetic agent which inhibits the binding of fibrinogen to activated platelets, and platelet aggregation. 3H-SR121566 bound with nanomolar affinity (KD ranging from 45 to 72 nM) to Gp IIb-IIIa expressing cells only. On activated human platelets, this ligand allowed the detection of a maximal number of 100-140,000 binding sites. The binding of SR121566 to platelets, was displaced by several agents including RGD-containing peptides and synthetic RGD mimetics, but not by ReoPro®, a humanised monoclonal antibody which inhibits the binding of fibrinogen to the Gp IIb-IIIa complex. Neither the fibrinogen dodecapeptide nor fibrinogen itself were able to compete with SR121566 whether platelets were activated or not.Flow cytometry studies indicated that SR121566 which did not activate Gp IIb-IIIa by itself, dose-dependently prevented the detection of activation-induced binding sites on TRAP-stimulated platelets in the presence or absence of exogenous fibrinogen, indicating a direct effect on the activation state of the Gp IIb-IIIa complex. Moreover, SR121566 was able to reverse the activation of Gp IIb-IIIa and to displace the binding of fibrinogen when added up to 5 min after TRAP stimulation of platelets. When added at later times (15 to 30 min), SR121566 failed to displace fibrinogen binding, even if SR121566 binding sites were still accessible and the Gp IIb-IIIa complex not activated.In conclusion, our study is in accordance with the finding that fibrinogen is recognised by the activated Gp IIb-IIIa complex through the dodecapeptide sequence present on its gamma chain, and that this interaction is inhibited by SR121566 by preventing and reversing the activated conformation of Gp IIb-IIIa and not by direct competition with fibrinogen.

scholarly journals Ligand-binding properties of proalbumin Christchurch

1980 ◽  
Vol 191 (1) ◽  
pp. 281-283 ◽  
Author(s):  
R G Reed ◽  
T Peters ◽  
S O Brennan ◽  
R W Carrell
Keyword(s):  
Fatty Acids ◽  
Human Serum Albumin ◽  
Ligand Binding ◽  
Serum Albumin ◽  
Human Serum ◽  
Protein Conformation ◽  
Binding Sites ◽  
Bromocresol Green ◽  
Binding Properties ◽  
Normal Albumin

Proalbumin Christchurch, a circulating variant of human serum albumin, is secreted from the liver without cleavage of the hexapeptide situated at the N-terminal end of the peptide chain of proalbumin. We compared ligand-binding properties of proalbumin Christchurch and of normal albumin A from the same individual in order to test the effect of the presence of the hexapeptide. The two albumin forms exhibited similar affinities for palmitate, bilirubin, 8-anilinonaphthalene-1-sulphonate and Bromocresol Green. The patterns of endogenous fatty acids bound to the two forms of albumin were slightly different, although the differences were probably not of physiological significance. From these studies it would appear that the propeptide of proalbumin does not alter the protein conformation in such a way as to alter binding sites for organic anions.

Relationship Between Aggregation and Binding of 125I-Fibrinogen and 45Calcium to Human Platelets

1979 ◽  
Author(s):  
E.I. Peerschke ◽  
R.A. Grant ◽  
M.B. Zucker
Keyword(s):  
Binding Sites ◽  
Calcium Binding ◽  
Silicone Oil ◽  
Human Platelets ◽  
Mechanisms Of Action ◽  
Ionized Calcium ◽  
Clot Retraction ◽  
Fibrinogen Binding ◽  
Induced Aggregation ◽  
Stimulation Of

Since calcium and fibrinogen are essential cofactors for ADP-induced aggregation, their mechanisms of action were investigated. Aspirin-treated platelets were filtered through Sepharose 2B equilibrated with cation-poor Tyrode’s solution. After adding the radioactive compounds at 22, platelets were centrifuged through silicone oil. Trapping was assessed in separate samples with 14C-sorbitoi. Calcium binding was maximal at 1 hr and with 200 uH CaCl2. Two binding sites could be demonstrated on normal and thrombasthenic platelets and on platelets which had lost their ability to aggregate (but not to change shape or promote clot retraction) after treatment with EDTA (8 min, 37°, pH 7.8). ADP did not alter calcium binding in the presence or absence of fibrinogen. Fibrinogen, however, bound to normal gel filtered platelets in the presence of ADP and ionized calcium “ but not to thrombasthenic or EDTA-treated platelets or to normal platelets in the presence of EDTA or at pH 6.5. Binding of fibrinogen increased with concentration but saturation was not observed even at physiologic levels. Fibrinogen binding was similar in gel filtered platelets and citrated piatelet-rich plasma. These studies indicate that stimulation of platelets with ADP under conditions favorable to aggregation is associated with binding of fibrinogen but not of additional calcium.

BINDING OF 37-DKa PLATELET AGGLUTINATING PROTEIN TO HUMAN PLATELETS

1987 ◽  
Author(s):  
E C -Y Lian ◽  
F A Siddigui
Keyword(s):  
Binding Sites ◽  
Polyclonal Antibodies ◽  
Helix Pomatia ◽  
Direct Interaction ◽  
Normal Subjects ◽  
Human Platelets ◽  
Scatchard Analysis ◽  
Specific Stimulation ◽  
Helix Pomatia Lectin ◽  
Stimulation Of

We have previously reported the purification of a 37-KDa platelet agglutinating protein (PAP p37) from the plasma of a patient with thrombotic thrombocytopenic purpura. using 125I-labeled p37, the properties of its binding to platelets were studied. The binding of p37 to washed human platelets from 4 normal subjects and two TTP patients after recovery was specific, concentration dependent and saturable. The Scatchard analysis revealed that the binding sites for p37 was about 100,000 per platelet with a dissociation constant of 48 × 10−9 M. The binding of p37 to erythrocytes was very little and non-specific. Stimulation of platelets by thrombin or ADP did not have any effect on the binding of p37 to platelets. The monoclonal antibodies to platelet GP lb (6D1) and GP Ilb-llla (10E5)(A gift of Dr. Barry coller) did not inhibit the binding of p37 to platelets. Fibrinogen (1 mg/ml) and FVIII/vWF (250 ug/ml) reduced the binding slightly. The polyclonal antibodies to p37 as well as concanavalin-A inhibited the binding of p37 to platelets through their direct interaction with p37. Other lectins such as phytohemagglutinin, potato lectin and helix pomatia lectin did not have any effect. At 40 mM, sialic acid, α-D-(+)-glucose, D-(+)-mannose and D-fructose caused 91%,44%,79%, and 63% inhibition of p37 binding respectively. D-(+)-galactose did not interfere with the binding. It is concluded that p37 binds to platelets on the sites other than GP lb and Gp IIb-IIIa and its binding to platelets is inhibited by certain sugars.

scholarly journals Mitogenicity and binding properties of β-galactoside-binding lectin from chick-embryo kidney

1981 ◽  
Vol 195 (2) ◽  
pp. 435-439 ◽  
Author(s):  
M J Pitts ◽  
D C H Yang
Keyword(s):  
Lymph Node ◽  
Chick Embryo ◽  
Binding Sites ◽  
Thymidine Incorporation ◽  
Mitogenic Activity ◽  
Binding Properties ◽  
Binding Lectin ◽  
Stimulation Of

THe beta-galactoside-binding lectin binds to glucosamine, mannosamine and galactosamine in addition to beta-galactoside, as determined by the inhibition of haemagglutination. Haemagglutination is further extended to examine the interaction of the binding sites for hexosamines and beta-galactosides, indicating that the binding of hexosamine and beta-galactoside is competitive. The lectin also shows strong mitogenic activity toward lymphocytes from mouse lymph node, as determined by the stimulation of thymidine incorporation.

scholarly journals NKG2D: Binding Properties for Glycan Ligands, and Mutagenesis Analysis

2011 ◽  
Vol 5 (1) ◽  
pp. 33-38
Author(s):  
Koji Higai ◽  
Sayo Matsumoto ◽  
Megumi Kimura ◽  
Yuzo Imaizumi ◽  
Kazuyuki Yanai ◽  
...  
Keyword(s):  
Bovine Serum Albumin ◽  
Ligand Binding ◽  
Binding Sites ◽  
Bovine Serum ◽  
Hepg2 Cells ◽  
Sialyl Lewis X ◽  
Binding Properties ◽  
Sialyl Lewis ◽  
Ligand Binding Sites ◽  
Class 1

Killer lectin-like receptor NKG2D, which is found on natural killer cells, recognizes MHC class 1-related ligands and also interacts with glycan ligands, heparin-conjugated bovine serum albumin (heparin-BSA) and sialyl Lewis X (sLeX) on multi-antennary N-glycans on transferrin secreted by HepG2 cells (HepTF). Using the glutathione-Stransferase- fused extracellular domain (AA 73-216) of NKG2D (rGST-NKG2D) and seven site-directed mutants, we explored in detail the binding of NKG2D to sulfate-containing glycan-BSA and HepTF. rGST-NKG2D binds to sulfatecontaining glycan-BSA with Kd values of 25 nM ±15 for λ-carrageenan-BSA, 66 ±23 nM for fucoidan-BSA, and 1.5±0.5 μM for heparan sulfate-BSA. Of the site-directed rGST-NKG2D mutants, Y152A, Q185A, K197A, Y199A, E201A, and N207A reduced binding to these glycans. These results indicate that NKG2D interacts with highly sulfated- and α2,3- NeuAc-containing glycans and suggest that the glycan-binding sites on NKG2D are shared between sulfate- and α2,3- NeuAc-containing glycans, and might overlap with protein ligand binding sites.

Colchicine Uptake and Binding by Human Platelets

1975 ◽  
Vol 34 (03) ◽  
pp. 780-794 ◽  
Author(s):  
Dianne M Kenney ◽  
Francis C Chao ◽  
James L Tullis ◽  
Gail S Conneely
Keyword(s):  
Time Dependence ◽  
Incubation Period ◽  
Binding Sites ◽  
Silicone Oil ◽  
Cold Treatment ◽  
Human Platelets ◽  
Temperature Dependent

SummaryThe uptake and binding of antimitotic alkaloid colchicine has been demonstrated in washed preparations of human platelets. A silicone oil technique was adapted so that both uptake and binding of 14C-colchicine were examined in the same platelet preparations. The time dependence and amount of colchicine taken up and bound by different platelet preparations during a 90 to 120 min incubation period were highly reproducible. Both colchicine uptake and binding by intact platelets, and colchicine binding by preparations of lysed platelets were specific and temperature dependent. Colchicine uptake was slowly reversible. Magnesium and GTP enhanced colchicine binding by lysed platelet preparations but calcium decreased binding.Exposure of platelets to either cold (4° C) or to thrombin, which disrupt platelet microtubules, produced significant increases in colchicine uptake and binding. The thrombin effect was maximal at 37° C and resulted in a greater increase in uptake and binding than that produced by either cold treatment alone or, by cold treatment followed by incubation with thrombin at 37° C. The amount of increase in uptake and binding produced by thrombin was independent of both thrombin (1–5 Units/109 platelets) and colchicine concentrations (1–50 × 10−6M).It is postulated that thrombin may initiate the formation, or make available, colchicine binding sites (microtubule subunits) within platelets.

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