Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q9UIJ5 (Rec)
 58,342 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The relationship between thrombomodulin-associated O-linked glycosammoglycans (GAGs) and the exogenous GAGs heparin or dermatan sulfate was studied in the inhibition of thrombin by antithrombin III (AT III) or heparin cofactor II (HC II). Both rabbit thrombomodulin (TM) and two glycoforms (a high-Mr form containing GAGs and a low-Mr form lacking the majority of O-linked GAGs) of a recombinant human TM deletion mutant (rec-TM) were used. The rapid inactivation of thrombin by HC II in the presence of dermatan sulfate was prevented by both the high-Mr rec-TM and the rabbit TM. In contrast, both rabbit TM treated with chondroitin ABC lyase to remove O-linked GAGs and the low-Mr form of rec-TM had only weak protecting effects. In the absence of exogeneous dermatan sulfate, thrombin inhibition by a high concentration of HC II was slightly accelerated by the high-Mr form of rec-TM but protected by rabbit TM. When thrombin inhibition by AT III in the presence of heparin was studied, both high-Mr rec-TM and rabbit TM again invoked a similar reduction of inactivation rates, whereas in the absence of exogenous heparin, both high-Mr forms accelerated thrombin inhibition by AT III. The diverse reactivities of various forms of TM towards HC II and AT III were also observed during protein C activation by the thrombin-TM complex. These results suggest that thrombin activity at the vessel wall or in fluid phase may undergo major kinetic modulations depending on the type of protease inhibitor, the presence or absence of exogenous GAGs and the glycosylation phenotype of TM. The dependence of TM anticoagulant function on the presence of an intrinsic GAG moiety suggests that variant glycoforms of this endothelial cell cofactor may be expressed differently in a species-, organ-, or tissue-specific manner as a means to regulate TM function in diverse vasculatures.
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PMID:Different glycoforms of human thrombomodulin. Their glycosaminoglycan-dependent modulatory effects on thrombin inactivation by heparin cofactor II and antithrombin III. 164 16

Two glycoforms of a soluble mutant of recombinant human thrombomodulin (rec.TM) were used to identify critical N- and O-linked glycans of the endothelial cell thrombin receptor. While N-linked glycans were not found to be involved in any function of rec.TM, an acidic chondroitin sulphate-like glycosaminoglycan (CSGAG) was found to be critical for all the direct anticoagulant functions of rec.TM, including inhibition of thrombin-mediated platelet aggregation. A glycoform of rec.TM lacking CSGAG had very poor anticoagulant activity. Furthermore, the glycoform of rec.TM possessing CSGAG showed strong inhibition by and had high affinity for poly-cationic basic proteins, whereas the CSGAG-deficient rec.TM did not. Monoclonal antibody binding as well as lectin mapping of rec.TM with agglutinins identified sialic acid containing O-linked glycans in both glycoforms additional to the CSGAG in high molecular weight rec.TM These findings define important molecular interactions modulating the anticoagulant function of TM, which appear to be critically regulated by CSGAG, and also showed that the overall post-translational glycosylation pattern of the two glycoforms was very similar except for the presence of CSGAG. The possibility exists that differently expressed glycoforms of TM may be crucial for the expression of endothelial cell-related anticoagulant potential in different vascular beds.
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PMID:Relationship between post-translational glycosylation and anticoagulant function of secretable recombinant mutants of human thrombomodulin. 165 91

Actin polymerization is an essential component of platelet activation. Since actin appears to polymerize at its membrane-associated end, knowledge of the structural relationship of actin filaments to membrane is an important part of understanding that polymerization process. A membrane-associated actin-containing cytoskeleton has been described in human platelets biochemically and is composed, at least in part, by an association between glycoprotein Ib and the actin-binding protein originally isolated from macrophages. Many other actin-associated proteins with known sub-membranous localization in other systems have been found in platelets, including alpha-actinin, vinculin, and low levels of spectrin and the red cell protein Band 4.1. Because of the density of the platelet cytoplasm, the structure of the membrane-skeleton has not yet been visualized. We have used quick freeze-deep etch techniques to observe the sub-membranous cytoplasm and report visualization of a periodic, submembranous filament system not before seen in the platelet. This filamentous system was more easily observed in thrombin-stimulated platelets, but appeared to be present in resting, discoid cells as well. The filaments could also be readily observed when platelets are lysed after fixation, stained with tannic acid, and embedded for thin-sectioning. This membrane cytoskeleton was composed of 9 nm thick filaments lying 15 nm apart, and 15 nm from the membrane. The filaments appeared to lie in parallel and to encircle the cell. Similar filaments could be seen associated with intracytoplasmic membrane systems in activated cells.
Anat Rec 1990 May
PMID:Platelet membrane skeleton revealed by quick-freeze deep-etch. 236 21

Blood coagulation tests were performed on dairy cattle in a herd with haemorrhagic problems on a farm in Gloucestershire. The characteristic pattern of prolonged partial thromboplastin time with normal prothrombin time and thrombin time was shown to be associated with a partial factor XI deficiency, a congenital defect previously identified in cattle in North America.
Vet Rec 1987 Jul 04
PMID:Identification of factor XI deficiency in Holstein-Friesian cattle in Britain. 362 78

Urokinase-type plasminogen activator (uPA) binds with high affinity to a specific cell surface glycosyl phosphatidyl-inositol (GPI)-anchored receptor, the urokinase receptor (uPAR). Pro-uPA, the enzymatically inactive single-chain form of uPA after having been activated by certain proteases, converts plasminogen into plasmin. This activation of pro-uPA to enzymatically active uPA can be prevented by the action of thrombin on pro-uPA. This inactivation process is accelerated in the presence of thrombomodulin (TM). The present study investigated whether pro-uPA bound to uPAR is still susceptible to inactivation by thrombin in the presence or absence of TM. A truncated soluble form of the uPAR lacking the GPI-anchor was cloned and expressed in CHO-cells (rec-uPAR277). Rec-uPAR277 efficiently inhibited the thrombin-mediated inactivation of pro-uPA up to 90% in a concentration dependent manner. The protective effect of rec-uPAR277 was far less pronounced when thrombin was complexed with TM. Enzyme kinetic experiments with varying concentrations of pro-uPA showed that in the presence of TM the catalytic efficiency (kcat/Km) of thrombin-mediated inactivation raised from 0.010 microM-1 s-1 to 0.50 microM-1 s-1 corresponding to a fifty-fold increase. In the presence of rec-uPAR277, however, the catalytic efficiency dropped by 4.1-fold (0.5 microM-1 s-1 to 0.122 microM-1 s-1). The inactivation kinetics of pro-uPA by thrombin (no TM added) in the presence of an excess of rec-uPAR277 could not be determined since virtually no inactivation occurred. Our data suggest that pro-uPA once bound to uPAR, is significantly protected from inactivation by thrombin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Inactivation of receptor-bound pro-urokinase-type plasminogen activator (pro-uPA) by thrombin and thrombin/thrombomodulin complex. 784 Sep 2

Rats which were infected with the gramnegative pathogen Klebsiella pneumoniae develop disseminated intravascular coagulation (DIC), multi-organ failure (MOF) and finally die in a septic shock. We investigated the therapeutic effect of antibiotic (tobramycin) treatment combined with the infusion of the highly specific thrombin inhibitor rec. hirudin. Although administration of 2 mg/kg tobramycin alone leads to a decrease of the bacterial burden, DIC could not be prevented. Infusion of rec. hirudin (0.25 mg/kg x h) for 4 h (start of treatment 1 h post infection), in addition to a bolus administration of tobramycin, led to an amelioration of DIC parameters as fibrinogen, thrombin-antithrombin complex (TAT) and platelets. Serum transaminase levels (GOT, GPT) as a marker of MOF were significantly improved by rec. hirudin, the T50 value increased from 17 h in the tobramycin group to 42 h in the tobramycin+rec. hirudin group, mortality rates were 90% or 60%, respectively. Combination of heparin (100 U/kg x h) and tobramycin was not effective on survival.
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PMID:Combination of antibiotic treatment with the thrombin inhibitor recombinant hirudin for the therapy of experimental Klebsiella pneumoniae sepsis. 797 45

An experimental disseminated intravascular coagulation (DIC) was induced in female CD rats by the intravenous administration of living bacteria (9.5 x 10(7) cfu Klebsiella pneumoniae), sublethal (5 mg/kg) or lethal (50 mg/kg) lipopolysaccharide (LPS), or tissue factor (1.5 micrograms/kg i.v. bolus or 0.4 micrograms/kg x hr i.v. infusion). We used a new fibrin monomer (FM) assay to follow the course of DIC. FM were detected by their ability to stimulate the tissue-type (t-PA) plasminogen activator dependent conversion of plasminogen to plasmin by a chromogenic assay. Miniplasminogen was used instead of plasminogen to avoid interference of the assay by alpha 2-antiplasmin. As a marker of DIC, elevated levels of FM were observed with all DIC-inducing agents (plasma levels were up to 90 micrograms/ml). The kinetics of FM formation were similar to the course of thrombin-antithrombin III (TAT) levels (maximal plasma levels 70 ng/ml); however, in the bacterial infection group, both parameters rose after a lag phase of about 1 hr. A 4 hr infusion of the highly specific thrombin inhibitor recombinant (rec.) hirudin (0.125 mg/kg x hr) resulted in a decrease of FM levels from 89.2 +/- 14.4 micrograms/ml in the LPS group (n = 10) to 27.4 +/- 11.2 micrograms/ml in the rec. hirudin group (n = 10; P < 0.001). The respective values for TAT levels were 73.1 +/- 19.7 micrograms/ml in the LPS group and 52.7 +/- 15.7 ng/ml in the rec. hirudin group (P < 0.001). Other coagulation parameters, such as platelets, fibrinogen, and fibrin(ogen) degradation products, were ameliorated accordingly.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Formation of fibrin monomers in experimental disseminated intravascular coagulation and its inhibition by recombinant hirudin. 805 64

C1-Inhibitor (Berinert, C1 INH), a 104 kDa protein, inhibits complement components (C1 esterase) as well as enzymes of the contact phase of coagulation (Factor XII, Factor XI) and kallikrein, thus regulating kinin generation. C1 INH is used for the treatment of the hereditary angioneurotic edema. This paper will give a survey about the evidence in recent literature concerning the potential efficacy of the compound on other diseases associated with shock, capillary leakage and inflammation as well. In our own experiments we evaluated whether the compound could influence acute inflammatory reactions or the severe systemic inflammatory response syndrome (SIRS) as a consequence of an experimental septic shock. To prevent the sepsis-induced DIC we co-infused the thrombin inhibitors AT III or rec. hirudin in combination with C1 INH. Coinfusion of C1-inhibitor (50-200 U/kg x h) with either rec. hirudin or AT III significantly improved survival rate compared to thrombin inhibitor alone.
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PMID:Influence of C1-inhibitor on inflammation, edema and shock. 817 80

Thrombomodulin (TM), a membrane proteoglycan on endothelial cells, binds thrombin in a 1:1 complex, accelerates the protein C activation by thrombin, promotes the thrombin inactivation by antithrombin III and inhibits the procoagulant properties of thrombin. The inactivation of single-chain urokinase-type plasminogen activator (scu-PA) by thrombin is accelerated about 70-fold by TM [De Munk, Groeneveld and Rijken (1991) J. Clin. Invest. 88, 1680-1684]. The present study investigates the role of the O-linked glycosaminoglycan moiety of TM in the latter reaction. In the presence of an excess of a fully-glycosylated soluble recombinant human TM mutant (high-Mr rec-TM), 0.11 nM thrombin inactivated 50% of 4.4 nM scu-PA in 45 min at 37 degrees C. In the presence of a soluble recombinant TM mutant lacking the glycosaminoglycans (low-Mr rec-TM), 1.9 nM thrombin was needed to inactivate 50% scu-PA, as compared with 4.7 nM thrombin in the absence of TM. Using the scu-PA inactivation assay the dissociation constant for the thrombin-TM interaction was found to be 0.4 nM for high-Mr rec-TM and 14 nM for low-Mr rec-TM. Treatment of high-Mr rec-TM with chondroitinase ABC to digest the glycosaminoglycans decreased the accelerating effect to the level of low-Mr rec-TM. A similar decrease was observed after treatment of solubilized rabbit TM with chondroitinase ABC. As expected, chondroitinase ABC had no influence on the accelerating effect of low-Mr rec-TM. The free glycosaminoglycans obtained by alkaline treatment of TM or chondroitin sulphate A also accelerated the inactivation of scu-PA by thrombin, but about 1000-fold higher concentrations than with TM were needed to obtain the same acceleration. It is concluded that the major glycosaminoglycan of TM plays a pivotal role in the inactivation of scu-PA by the TM-thrombin complex, both in the formation and in the activity of the complex.
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PMID:Role of the glycosaminoglycan component of thrombomodulin in its acceleration of the inactivation of single-chain urokinase-type plasminogen activator by thrombin. 838 42

Coagulation factor XI is a glycoprotein of the contact factor system. Its deficiency is associated with a highly variable bleeding tendency, thus a role in relation to hemostasis appears to exist. However, the importance of factor XI for stimulating intrinsic coagulation in vivo has not yet been determined. To study the procoagulant effects of human factor XIa in vivo, we infused the purified enzyme into normal chimpanzees (100 micrograms) in the absence or presence of the thrombin inhibitor rec-hirudin (1.0 mg/kg loading dose plus 0.3 mg/kg body wt continuous infusion). Factor XIa elicited an immediate activation of factors IX, X, and prothrombin, as measured by their respective activation fragments. However, whereas the activation of factors IX and X was immediate and shortlasting, (peak increments of 6- and 1.4-fold of baseline at 5 minutes after injection), the conversion of prothrombin gradually increased, reaching a summit of 6-fold baseline values after 60 min, and remaining elevated during the course of the experiments. Thrombin-antithrombin complexes also remained elevated during the study period. In the presence of hirudin, the initial activation of factors IX, X, and prothrombin was unchanged, however the further increment in prothrombin fragment F1 + 2 was markedly inhibited. These results demonstrate that factor XIa is a potential agonist of the intrinsic cascade in vivo, which activity is enhanced in the presence of thrombin.
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PMID:Factor XIa induced activation of the intrinsic cascade in vivo. 870 5


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