Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have examined initial assembly of the extrinsic pathway of blood coagulation on cell surfaces with radiolabeled human factor VIIa and a human fetal lung cell line possessing abundant functional tissue factor activity. Binding of factor VIIa to these cells was observed and was time- and temperature-dependent. Binding of factor VIIa was quantitatively equivalent at 37 and 6 degrees C, although the kinetics of binding differed. The radiolabeled ligand bound by the cell was indistinguishable by sodium dodecyl sulfate-polyacrylamide gel analysis from the factor VIIa offered. Factor VIIa binding was influenced by calcium ions. The binding appears to involve at least two classes of calcium-dependent binding sites. Optimal binding occurred at 2 mM calcium for both classes of sites, and there was inhibition of binding to the high affinity sites at higher calcium. Association of factor VIIa was specific, saturable, had a Kd of 123 +/- 37 pm, and factor VIIa interacted with about 100,000 binding sites per cell. Once established, specific binding was rapidly reversible. Direct cellular binding of human factor X also was observed and was calcium, time- and temperature-dependent. Factor X binding was specific and saturable with half-maximal binding at 87.6 +/- 27.4 nM to 6.03 +/- 1.03 X 10(6) sites per cell. Specific high affinity binding of factor VIIa correlated with generation of factor Xa. A direct linear relationship was observed at low factor VIIa binding; however, at higher bound factor VIIa, the relationship was nonstoichiometric, i.e. less factor Xa was formed per mole of factor VIIa. Expression of specific binding sites for factors VIIa and X provides further substantiation for the molecular assembly hypothesized to initiate the extrinsic coagulation protease cascade on cells.
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PMID:Initiation of the extrinsic pathway of coagulation. Association of factor VIIa with a cell line expressing tissue factor. 349 35

The binding of factor VII and tissue factor produces a membrane-associated proteolytic complex which may be the primary biological initiator of coagulation. Homogeneous tissue factor, a glycoprotein purified from bovine brain, was reconstituted into phospholipid vesicles ranging from neutral (100% phosphatidylcholine) to highly charged (40% phosphatidylserine) with octyl glucoside. The vesicles were characterized with respect to size and tissue factor content and orientation. Employing data from protease digestion, we deduced that tissue factor is randomly oriented; thus, its effective concentration in these vesicles was half its total concentration. In all binding experiments, 1 mol of enzyme was bound per mole of available activator at saturation. This stoichiometry was not affected by the form of the enzyme employed or the phospholipid composition of the vesicles. With tissue factor incorporated into phosphatidylcholine vesicles, the Kd was 13.2 +/- 0.72 nM for factor VII and 4.54 +/- 1.37 nM for factor VIIa. Thus, the one-chain zymogen binds to the activator with only slightly less affinity than the more active two-chain enzyme. Active-site modification of factor VII and factor VIIa with diisopropyl fluorophosphate resulted in tighter binding of the derivatized molecules. Inclusion of phosphatidylserine in the vesicles altered the binding both quantitatively and qualitatively. With increasing acidic phospholipid, the concentration of enzyme required to occupy half the activator sites was decreased. In addition, positive cooperativity was observed, the degree of which depended on the vesicle charge and the form of the enzyme. An explicit two-site cooperative binding model is presented which fits these complex data. In this model, tissue factor is at least a dimer with two interacting enzyme binding sites.
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PMID:Factor VII binding to tissue factor in reconstituted phospholipid vesicles: induction of cooperativity by phosphatidylserine. 352 61

The kinetics of activation of human Factor IX by human Factor XIa was studied by measuring the release of a trichloroacetic acid-soluble tritium-labeled activation peptide from Factor IX by a modification of a method described for bovine Factor IX activation by Zur and Nemerson (Zur, M., and Y. Nemerson, 1980, J. Biol. Chem., 255:5703-5707). Initial rates of trichloroacetic acid-soluble 3H-release were linear over 10-30 min of incubation of Factor IX (88 nM) with CaCl2 (5 mM) and with pure (greater than 98%) Factor XIa (0.06-1.3 nM), which was prepared by incubating human Factor XI with bovine Factor XIIa. Release of 3H preceded the appearance of Factor IXa activity, and the percentage of 3H released remained constant when the mole fraction of 3H-labeled and unlabeled Factor IX was varied and the total Factor IX concentration remained constant. A linear correlation (r greater than 0.98, P less than 0.001) was observed between initial rates of 3H-release and the concentration of Factor XIa, measured by chromogenic assay and by radioimmunoassay and added at a Factor IX:Factor XIa molar ratio of 70-5,600. Kinetic parameters, determined by Lineweaver-Burk analysis, include Km (0.49 microM) of about five- to sixfold higher than the plasma Factor IX concentration, which could therefore regulate the reaction. The catalytic constant (kcat) (7.7/s) is approximately 20-50 times higher than that reported by Zur and Nemerson (Zur, M., and Y. Nemerson, 1980, J. Biol. Chem., 255:5703-5707) for Factor IX activation by Factor VIIa plus tissue factor. Therefore, depending on the relative amounts of Factor XIa and Factor VIIa generated in vivo and other factors which may influence reaction rates, these kinetic parameters provide part of the information required for assessing the relative contributions of the intrinsic and extrinsic pathways to Factor IX activation, and suggest that the Factor XIa catalyzed reaction is physiologically significant.
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PMID:Kinetics of the Factor XIa catalyzed activation of human blood coagulation Factor IX. 660 71

Not only does tissue factor (TF) play a crucial role in hemostasis and thrombosis, but it is also involved in tumor progression and metastatic potency in some malignant tumors. We evaluated the clinical relevance of TF expression in melanocytic tumors and TF serum level in patients with malignant melanoma. TF expression in benign and malignant melanocytic lesions was examined by immunoperoxidase staining in 20 nevi, 41 primary, and 24 metastatic melanoma lesions. TF was detected in 94, 95, and 100% of these lesions, respectively. The staining pattern was membranous and cytoplasmic both in nevi and melanoma cells. This finding was confirmed by western blot analysis using cultured human melanocytes, nevi cells, and melanoma cell lines. TF was also expressed on blood vessels in benign and malignant melanocytic lesions. Expression of TF in primary melanoma lesions was not associated with any clinicopathological variables. In addition, the serum level of TF was elevated in 14% of patients with melanoma; however, it was not correlated with disease progression. These results suggest that TF was ubiquitously expressed in melanocytic cells and its expression was not correlated with disease progression and/or metastatic potency of melanoma cells.
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PMID:Tissue factor expression and serum level in patients with melanoma does not correlate with disease progression. 1143 67

Tissue factor (TF) is involved in tumor progression and metastatic potency in some malignant tumors and its function is regulated by tissue factor pathway inhibitor (TFPI) therefore the interaction of both molecules is crucial for their functional role. We evaluated the clinical relevance of TF and TFPI expression in benign and malignant melanocytic lesions. Expression of both was examined by immunoperoxidase staining using serial tissue sections in 16 nevi, 34 primary and 15 metastatic melanoma lesions. TF and TFPI were ubiquitously expressed in benign and malignant melanocytic lesions. This finding was confirmed by Western blot analysis using cultured human melanocytes, nevi cells (NCN) and melanoma cell lines. Although TF expression was not associated with malignant transformation and disease progression, TFPI expression in primary and metastatic melanoma lesions was significantly lower and weaker than that in nevi lesions in terms of intensity and percentage of stained cells. In addition, TFPI expression in metastatic lesions was significantly lower and weaker than that of TF. These results suggest that the relative expression of TF to TFPI may play a crucial role in the malignant transformation and metastatic potency in melanocytic cells.
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PMID:Differential expression of tissue factor and tissue factor pathway inhibitor in metastatic melanoma lesions. 1202 85

Derivatives containing arginine-glycine-aspartic acid (RGD) inhibit fibrinogen binding to activated platelets and promote endothelial and smooth muscle cell attachment. An amphiphilic derivative of RGD that can be dissolved in an organic solvent has potential in the development of non-thrombogenic biomaterials. Such a derivative, LA-GRGD, was synthesised by coupling glycine-arginine-glycine-aspartic acid (GRGD) with lauric acid (LA). Its solubility and antithrombotic, cytotoxic and cell-binding effects were then evaluated in comparison with heparin (which is used clinically) and a fibronectin-engineered protein polymer (FEPP). Thromboelastography (TEG) was used to measure blood clotting time using fresh whole blood from healthy volunteers. Tissue factor (TF) activity was measured using plasma with a standard prothrombin time assay (PT). Cytotoxicity was assessed on human umbilical cord endothelial cells (HUVECs) using an Alamar blue assay. Solubility of the conjugate was assessed in a co-solvent. These techniques were used to study LA-GRGD, using heparin and FEPP as controls. The amphiphilic property of LA-GRGD was dependent on the feed mole ratio of GRGD to LA. LA-GRGD was soluble in acetone:water and water. LA-GRGD inhibited TF by >90% and prolonged TEG-r by 8.2+/-3.3 min (200 microg ml(-1)). Heparin inhibited TF by >90%, but prolonged TEG-r by 97.4+/-1.6 min (1 U ml(-1)); FEPP inhibited TF by >90% (100 microg ml(-1)) and prolonged TEG-r by 73.7+/-8.4 min (10 microg ml(-1)). Heparin had no cytotoxic effect on EC metabolism and viability at the concentrations studied (0.1-100 U ml(-1)). No significant cytotoxic effect was produced by LA-GRGD or FEPP at concentrations ranging from 0.1 microg ml(-1) to 50 microg ml(-1), but, at higher concentrations (100 microg ml(-1) and 200 microg ml(-1)), a detrimental effect was observed. Cell binding studies showed that LA-GRGD bound 29% of ECs compared with FEPP (60%) and heparin (22%). This new approach for synthesising amphiphilic RGD and its analogues has potential as a drug delivery system for the manufacture of new polymer formulations for use in bypass grafts and other tissue-engineered devices.
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PMID:Synthesis and evaluation of amphiphilic RGD derivatives: uses for solvent casting in polymers and tissue engineering applications. 1468 1