Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To investigate mechanisms regulating intra-alveolar coagulation, we studied monolayers of the A549 human lung epithelial cell line. The surface of A549 cells delayed the onset of prothrombin-to-thrombin conversion and prevented total prothrombin consumption in normal plasma compared to plastic cell-free wells. Similar results were achieved with bovine pulmonary endothelial (CPAE) and rat intestinal epithelial (IEC-6) cell lines, whereas Madin-Darby canine kidney renal epithelial cell line accelerated thrombin formation. The A549 surface catalyzed antithrombin III-thrombin complex formation with no significant increase in thrombin inactivation from heparin cofactor II. The A549 cell surface effects were largely, but not completely, reversed to values obtained for plastic when protein C-deficient plasma was used. Pretreatment of the cell surface with chondroitinase ABC plus heparitinase prior to thrombin generation experiments had no effect on the total prothrombin consumed but decreased the initial delay. Heparan sulfate as well as dermatan sulfate and other chondroitin sulfates were detected on the A549 surface using alcian blue staining. Conditioned media from A549, CPAE, and IEC-6 cells delayed the clot time of recalcified plasma. Use of chondroitinase ABC and heparitinase were both required to obliterate the A549 conditioned media activity. After growing A549 cells in 35SO(2-)4-containing medium, the resultant conditioned medium was found to contain 2,000 kD and 300- to 1,000-kD proteoglycans that yielded chains of less than or equal to 100 kD on reductive elimination with base.(ABSTRACT TRUNCATED AT 250 WORDS)
Am J Respir Cell Mol Biol 1991 Apr
PMID:A549 lung epithelial cells synthesize anticoagulant molecules on the cell surface and matrix and in conditioned media. 201

An isoflavone compound, genistein, which is known as a protein tyrosine kinase inhibitor, concentration-dependently (0.1-30 micrograms/ml) suppressed human platelet aggregation, serotonin secretion, and protein tyrosine phosphorylation induced by collagen or stable thromboxane A2 analogs [U46619 and 9,11-epithio-11,12-methano-thromboxane A2 (STA2)]. However, genistein did not inhibit these thrombin (0.1 unit/ml)-induced platelet responses. Although thrombin induced an increase in the platelet phosphotyrosine content, genistein at 100 micrograms/ml only slightly attenuated thrombin-induced protein tyrosine phosphorylation. Genistein competitively inhibited [3H]U46619 binding to washed platelets, in a concentration-dependent fashion. Daidzein (another isoflavone compound), which does not have a hydroxyl group at the 5-position of genistein and lacks inhibitory activity for protein tyrosine kinase, was found to suppress [3H]U46619 binding, leading to the inhibition of collagen- or STA2-induced platelet responses. These results indicate that the blockage by genistein of platelet responses induced by collagen or thromboxane A2 is due to its preventive action on thromboxane A2 binding to the receptor, rather than via inhibition of protein tyrosine phosphorylation, and that the drug does not appear to be a particularly good inhibitor of tyrosine phosphorylation in intact platelets.
Mol Pharmacol 1991 Apr
PMID:Genistein, a protein tyrosine kinase inhibitor, inhibits thromboxane A2-mediated human platelet responses. 201 48

Fibrinogen, the principal blood-clotting protein, is made up of three different subunits synthesized in the liver. In vitro administration of glucocorticoids to liver cells from the frog Xenopus laevis causes a dramatic increase in fibrinogen synthesis. Investigations of molecular mechanisms underlying this hormonal stimulation at the mRNA level require cDNA clones complementary to the mRNAs coding for the three fibrinogen subunits, called A alpha, B beta, and gamma. We describe here the isolation and characterization of cDNA clones for the B beta subunit of Xenopus fibrinogen. cDNA libraries in both plasmid (pBR322) and phage (lambda gt10) cloning vectors were constructed from frog liver mRNA and screened with a rat B beta cDNA. Clones thus isolated hybridized to two Xenopus liver mRNAs 2500 and 1800 bases long, the previously-determined sizes for B beta mRNAs. The identity of the plasmid clone B beta-27 was confirmed by hybridization-selection of complementary mRNA which translated in vitro into the B beta polypeptide, as determined by size and susceptibility to thrombin cleavage. lambda/B beta 10, a clone representing nearly all of the 2500-base B beta mRNA, was isolated from the phage cDNA library. The 3'-end of this clone includes a polyadenylation signal about 20 residues upstream of a stretch of 34 adenosine residues, which probably represents the 3'-poly(A) tail of the messenger RNA. lambda/B beta 10 lacks only 20 nucleotides of full-length B beta mRNA at the 5'-end and there is one major start site of transcription. The 2500-base B beta mRNA has a 700-base extension at the 3'-end that is not present in the 1800-base mRNA. The Xenopus laevis genome contains two or three genes for the B beta fibrinogen subunit. Using the cDNA clone as a probe, B beta mRNA was shown to be induced at least 20-fold by glucocorticoid treatment of purified parenchymal cells of Xenopus liver maintained in primary culture.
Mol Cell Endocrinol 1991 Feb
PMID:Molecular cloning of cDNA for the B beta subunit of Xenopus fibrinogen, the product of a coordinately-regulated gene family. 205 Feb 71

In this study we extracted glycoconjugates from bovine sublingual gland and attempted to investigate anticoagulant properties on blood coagulation of humans. To this purpose we performed the thromboelastographic technique and selective hemocoagulation screening tests. In particular, findings from screening tests indicated the occurrence of hemostatic effects on thrombin time (TT) and activated partial thromboplastin time (aPTT) with a response in a dose-related manner, whereas significant changes were not found on prothrombin time (PT).
Cell Mol Biol 1990
PMID:Anticoagulant activity with dose-related response of glycoconjugates from bovine sublingual gland. 207 80

The cDNA encoding bovine opsin was transfected into Chinese hamster ovary (CHO) cells to generate stable clones expressing the rod cell photoreceptor protein. Cells expressing opsin, when incubated in 11-cis retinal and exposed to light, inhibited forskolin-stimulated adenylyl cyclase activity. Rhodopsin-mediated inhibition of adenylyl cyclase was prevented by treatment of cells with pertussis toxin. In the same cells, thrombin stimulated phosphatidylinositol hydrolysis through G protein-mediated pathways, but rhodopsin neither significantly influenced the action of thrombin nor stimulated phosphatidylinositol hydrolysis. Our findings indicate that rhodopsin selectively regulates a Gi protein in intact CHO cells that is coupled to adenylyl cyclase but not to phospholipase C.
J Mol Endocrinol 1990 Feb
PMID:Rhodopsin expressed in Chinese hamster ovary cells regulates adenylyl cyclase activity. 210 93

Recent studies have shown that ligand-activated growth factor receptors as well as transforming versions of nonreceptor protein-tyrosine kinases physically associate with phosphatidylinositol-3 kinase (PI-3 kinase). Reasoning that PI-3 kinase might also play a role in the normal functions of nonreceptor kinases, we sought to determine whether association with PI-3 kinase might serve as a measure of nonreceptor protein-tyrosine kinase activation under physiological conditions. We found that p60c-src as well as p59fyn, the product of another member of the src family of proto-oncogenes, physically associated with a PI kinase activity within 5 s after exposure to thrombin. Furthermore, PI kinase reaction products generated in p60v-src, p60c-src or p59fyn containing immunoprecipitates were indistinguishable, demonstrating the identity of the associated enzyme as PI-3 kinase. These findings demonstrate a thrombin-dependent interaction between p60c-src or p59fyn and PI-3 kinase and suggest a role for nonreceptor protein-tyrosine kinases in human platelet signal transduction.
Mol Cell Biol 1990 Jul
PMID:Thrombin-dependent association of phosphatidylinositol-3 kinase with p60c-src and p59fyn in human platelets. 216 81

We investigated Plasmodium falciparum parasitized erythrocyte binding to proteolytic fragments of thrombospondin and the effects of anti-thrombospondin monoclonal antibodies on this binding. Purified human platelet thrombospondin was cleaved by trypsin, chymotrypsin or thrombin. Fragments were separated by heparin-agarose affinity chromatography, removing the amino-terminal heparin-binding region. Trypsin at 5.0 micrograms ml-1 of thrombospondin cleaved thrombospondin to reduced 140 and 120 kDa fragments plus a reduced 25-kDa heparin-binding fragment. Infected erythrocytes bound to intact thrombospondin (3420 +/- 460 infected erythrocytes mm-2) and the carboxy-terminal fragment, yielding 120-140-kDa fragments on sulfhydryl reduction, but not to the 25-kDa fragment (144 +/- 104 infected erythrocytes mm-2 (mean +/- s.d., N = 4). Similar results were obtained with chymotrypsin and thrombin cleavage. When the anti-thrombospondin monoclonal antibody MA-I was added to immobilized thrombospondin prior to infected erythrocytes, adherence was inhibited by 99%. At the same concentration, MA-I inhibited adherence to C32 melanoma cells by only 35%. MA-I binds to a calcium-dependent structure at the C-terminal globular region of thrombospondin. Monoclonal antibody MA-II inhibited adherence to thrombospondin by 46%, while MA-III had no effect. These antibodies bind to the N-terminal globular region which includes the heparin-binding site and the segment connecting the two globular regions, respectively. The site(s) for infected erythrocyte binding on thrombospondin reside in the large, 140- or 120-kDa, proteolytic cleavage fragments, and not in the N-terminal heparin-binding region.
Mol Biochem Parasitol 1990 May
PMID:Falciparum malaria parasitized erythrocytes bind to a carboxy-terminal thrombospondin fragment and not the amino-terminal heparin-binding region. 219 22

Continuous or repeated injury of rabbit aortae by indwelling vascular catheters caused the deposition of platelets on the injured vessels and the formation of thrombi rich in platelets and fibrin at sites where flow was most disturbed and injury was most extensive. Incorporation of 51Cr platelets into the thrombi reached a maximum between 3 and 24 hr. The platelet-fibrin-rich thrombi remained reactive to circulating platelets for at least 14 days. Continuing reactivity of thrombi and the turnover of platelets in the thrombi were accompanied by an increase in the proportion of platelets that separated in the least dense fraction on Stractan density gradients. Platelet survival was also shortened (43.5 +/- 5.9 hr in animals with catheters, compared with 62.6 +/- 4.5 hr in animals with a sham operation), indicating that some platelets that had taken part in thrombus formation or had interacted with the injured vessel wall were rapidly cleared from the circulation. Platelets from rabbits that had had indwelling aortic catheters in place for 3 or 6 days survived significantly longer than those from animals with a sham operation upon injection of the platelets into normal animals; thus, continuous turnover of platelets on injured vessels and thrombi, and the clearance of altered platelets, leads to a population of younger platelets that survive longer. The continuing reactivity of thrombi may in part account for repeated occlusive episodes in vascular disease. The contribution of thrombin generation and fibrin formation to the platelet-rich thrombi is substantial and warrants the ongoing evaluation of treatment with a combination of anticoagulant and antiplatelet agents in arterial thrombosis and in thrombus formation on vascular catheters.
Exp Mol Pathol 1990 Dec
PMID:The relation among vessel injury, thrombus formation, and platelet survival in rabbits. 225 31

Structures formed during the early stages of clot formation have been produced in a controlled manner by polymerization of soluble fibrin monomers prepared from dissolved normal clots that had been formed upon addition of thrombin to fibrinogen. In agreement with other studies using different approaches, electron microscopy of negatively contrasted or rotary-shadowed specimens of these preparations reveal two-stranded protofibrils, as well as shorter oligomers and fibrin monomers. Individual fibrin molecules are similar in appearance to fibrinogen, suggesting that no large-scale changes in conformation occur on removal of the fibrinopeptides. Moreover, these micrographs show details of the protofibril structure not previously seen. The visualization of clear cross-over points of the filaments making up the protofibril indicate that these structures are twisted. Diffraction patterns of electron micrographs of both protofibrils and fibers and computer modeling of protofibrils also suggest that these structures are twisted but not precisely ordered.
J Mol Biol 1990 Dec 05
PMID:Electron microscope investigation of the early stages of fibrin assembly. Twisted protofibrils and fibers. 225 25

The amidase activity of human alpha-thrombin has been studied in the pH range 5.5 to 10, and at four different chloride concentrations from 5 mM to 1 M. The Michaelis-Menten constant, Km, shows a bell-shaped dependence over the pH range studied, with a minimum around pH 8. The pH dependence of the catalytic constant, kcat, shows multiple inflection points especially at low (less than 0.1 M) chloride concentrations, thereby implicating the existence of multiple catalytic forms of the enzyme. A general linkage scheme is proposed for the analysis of the effect of protons on thrombin amidase activity, and experimental data have globally been analysed over the entire pH range in terms of such a scheme. Four proton-linked ionizable groups seem to be involved in the control of thrombin amidase activity. Two of these groups change their pK value upon substrate binding to the enzyme and account for the pH dependence of Km. All four groups control the catalytic activity of the enzyme which decreases with increasing protonation. Chloride has little effect on Km, while kcat changes significantly at pH less than 8. This effect is due to an increased enzymatic activity of the highly protonated intermediates at high chloride concentrations, as well as to the pK shift of two proton-linked ionizable groups.
J Mol Biol 1990 Dec 20
PMID:Effect of protons on the amidase activity of human alpha-thrombin. Analysis in terms of a general linkage scheme. 226 57


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