Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It is now well established that some serine proteases, such as plasminogen activators and thrombin, as well as their inhibitors, have roles in the development of both central and peripheral nervous systems. We have shown that muscle plasminogen activators, activated after denervation, were able to digest some components of the muscle basement membrane. We have also shown that several inhibitors of serine proteases were concentrated at the neuromuscular junction. These are protease nexin, I, also called glia-derived nexin, protease nexin II, a secreted form of the beta-amyloid precursor protein (APP), and alpha 1-antichymotrypsine (ACT). These results leads us to propose a model in which serine proteases would favor plasticity of motor nerve endings during neuromuscular development. On the contrary, the inhibitors of serine proteases would act to provide and secure maintenance of the synaptic contact. A dysequilibrium between serine proteases and their inhibitors might underlie one or more motor neuron diseases.
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PMID:[Serine proteases and their inhibitors: their role in the differentiation in neuromuscular system]. 778 Jul 96

Protein C inhibitor (PCI) is a heparin-binding serine proteinase inhibitor (serpin) which is thought to be a physiological regulator of activated protein C (APC). The residues F353-R354-S355 (P2-P1-P1') constitute part of the reactive site loop of PCI with the R-S peptide bond being cleaved by the proteinase. Changing the reactive site P1 and P2 residues to those of either proteinase nexin-1, alpha 1-proteinase inhibitor or heparin cofactor II resulted in a decrease in inhibitory activity towards thrombin and APC. Changing the P2 residue F353-->P generated a rPCI which was a better thrombin inhibitor, but was 10-fold less active with APC. While these results support the concept that the P1 and P2 residues are important in the specificity of PCI, they suggest that the reactive site residues are not the only determinant of serpin specificity. Kinetic analysis of the rPCI variants was consistent with PCI operating by a mechanism similar to that proposed for other serpins. In this model an intermediary complex forms between inhibitor and proteinase that can proceed to either cleavage of the inhibitor as substrate or formation of an inactive complex.
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PMID:Reactive site mutants of recombinant protein C inhibitor. 781 27

Protease nexin-1 (PN-1) is a potent inhibitor of serine proteases, such as thrombin and plasminogen activators, which is secreted into the extracellular space. Since PN-1 is induced following lesion of the sciatic nerve, the effect of substances known to accumulate at the site of injury was examined in primary cultures of Schwann cells. Among the cytokines, growth factors, mitogens, neurotrophins, and neuroactive peptides analyzed, only angiotensin II (Ang II), calcitonin gene-related peptide (CGRP), and vasoactive intestinal peptide (VIP) were found to regulate the expression of PN-1 on Schwann cells. While Ang II and CGRP caused downregulation, VIP acted as a positive modulator of PN-1. Displacement of Ang II binding using the selective ligands losartan and CGP 42112 led to a severalfold increase of PN-1 protein and mRNA over basal levels, indicating that the observed effect was mediated by specific binding sites. Indeed, the presence of AT1 and AT2 angiotensin receptor subtypes was demonstrated in cultured Schwann cells as well as in the rat sciatic nerve. Moreover, the detection of angiotensinogen- and renin-mRNA in these cultures suggested an endogenous production of Ang II. This data identified one of the mechanisms regulating PN-1 synthesis. Altogether our results indicate that neuropeptides can differentially control the proteolytic activity of the microenvironment, providing new aspects of neuron-glia interactions in the intact tissue and following nerve injury.
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PMID:Regulation of protease nexin-1 expression in cultured Schwann cells is mediated by angiotensin II receptors. 782 77

Protease nexin-2/amyloid beta-protein precursor (PN-2/A beta PP) is a Kunitz-type protease inhibitor which has been shown to be a tight-binding inhibitor of enzymes, factors XIa and IXa (FIXa), suggesting a role for this protein in hemostasis. Since coagulant reactions are modulated on biologic surfaces, we investigated how 25:75 (mol/mol) phosphatidylserine/phosphatidylcholine vesicles (PSPC), thrombin-activated platelets, or umbilical vein endothelial cells influence inactivation of FIXa by PN-2/A beta PP. The Km of human or porcine FIXa activation of human factor X in the presence of PSPC, activated platelets, or endothelial cells in the absence or presence of thrombin-activated factor VIII (FVIIIa) was similar, (0.05-0.39 microM). The presence of FVIIIa increased the catalytic efficiency (kcat/Km ratio) of human and porcine factor IXa's activation of factor X 4952-406-fold, respectively. In the presence of PSPC, the Ki of human and porcine FIXa inhibition by PN-2/A beta PP was Ki = 1.9 x 10(-9) M and 5.8 x 10(-9) M, respectively. After the addition of FVIIIa to the reaction, the Ki for both human and porcine FIXa inhibition by PN-2/A beta PP on PSPC increased 13- and 4-fold to Ki = 2.5 x 10(-8) M and 2.4 x 10(-8) M, respectively. These Ki for inhibition of human FIXa on phospholipid vesicles by PN-2/A beta PP were similar when factor X activation was measured by chromogenic or activation peptide release assays. FVIIIa reduced the inhibition of FIXa by PN-2/A beta PP only in the presence of PSPC.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Factor IXa inhibition by protease nexin-2/amyloid beta-protein precursor on phospholipid vesicles and cell membranes. 782 67

Our article documents recent studies in the proteolytic processing of the Alzheimer's beta-amyloid protein precursor (beta-APP), as well as the role of thrombin and its potent inhibitor, protease nexin I in Alzheimer's disease (AD). Since synapse loss correlates best with cognitive decline in AD, we also present in detail, our model of synapse formation and elimination, reviewing recent findings related to the subject as well as our own original data. Recent exciting findings concerning the involvement of thrombin-like activity in synapse elimination, which we feel to be important in neural plasticity are also discussed.
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PMID:Serine proteases and their serpin inhibitors in Alzheimer's disease. 785 60

Recent studies have shown that serine protease inhibitors can be regulated in their activity, specificity, and location by glycoprotein or extracellular matrix (ECM) co-factors. Protease nexin-1 (PN-1) is a member of the serpin superfamily of serine protease inhibitors which can rapidly inhibit thrombin, urokinase, and plasmin. PN-1 binds tightly to and is regulated by the ECM. This interaction accelerates the inhibition of thrombin by PN-1 and blocks urokinase and plasmin inhibition by PN-1. Previous work showed that heparan sulfate proteoglycan is largely responsible for the acceleration of thrombin inhibition by PN-1. Our current studies were directed at identifying ECM component(s) that decreased the ability of PN-1 to inhibit urokinase and plasmin. These studies showed that collagen type IV decreased the formation of SDS-stable complexes between urokinase or plasmin and PN-1 without affecting formation of complexes between thrombin and PN-1. The second order rate constant for inhibition of urokinase by PN-1 was markedly decreased with increasing collagen type IV, whereas the second order rate constant for inhibition of thrombin by PN-1 was unaffected by addition of collagen type IV. Other ECM components (collagen type I, vitronectin, fibronectin, and heat-denatured collagen type IV) did not affect complex formation or the rate of inhibition of proteases by PN-1, indicating that these effects were specific to collagen type IV. Binding of PN-1 to immobilized collagen type IV was demonstrated using an enzyme-linked immunosorbent assay; the concentration of PN-1 necessary to obtain 50% saturation of the immobilized collagen type IV binding sites was approximately 15 nM. Collagen type IV was also copurified with PN-1 from fibroblast-conditioned medium. These results demonstrate a novel regulation of serpin specificity in which an ECM co-factor decreased the inhibition of certain proteases by the serpin without affecting the inhibition of its target protease.
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PMID:Regulation of protease nexin-1 target protease specificity by collagen type IV. 800 28

Recombinant rat glia-derived nexin was expressed in insect cells using the baculovirus system. The kinetics for the inhibition of thrombin by this recombinant material were indistinguishable from those observed with natural glia-derived nexin and recombinant nexin expressed in yeast. In addition, the dependence of the rate of inactivation on the concentration of heparin was similar for the three preparations. At the optimal heparin concentration, the association rate constant was 330-fold higher than that observed in the absence of heparin. A putative heparin-binding site is found in glia-derived nexin between residues 71 and 86; heparin-binding sites are found in homologous regions of antithrombin III and heparin cofactor II. Lysines in this region were mutated to glutamates, and the kinetics for the inhibition of thrombin by mutant proteins were determined. Concurrent mutation of all seven lysines in this region (residues 71, 74, 75, 78, 83, 84, and 86) did not affect the rate constant for the association of glia-derived nexin with thrombin in the absence of heparin, but it resulted in complete loss of the heparin acceleration of the rate of association. Mutations of residues 83, 84, and 86 together also caused a marked decrease in the acceleration by heparin of the reaction between glia-derived nexin and thrombin. These results support the hypothesis that the heparin-binding sites of glia-derived nexin, antithrombin III, and heparin cofactor II are found in homologous regions of the molecules. Heparin was also found to potentiate the ability of wild-type glia-derived nexin to inhibit the thrombin-induced retraction of neurites from neuroblastoma NB2a cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Localization of the heparin-binding site of glia-derived nexin/protease nexin-1 by site-directed mutagenesis. 801 37

After binding to its receptor (uPAR), active cell-surface urokinase (uPA) is not internalized while the complex formed by uPA with plasminogen activator inhibitor type 1 (PAI-1) is internalized and degraded. Internalization and degradation require binding to uPAR and subsequently an interaction with the alpha 2-macroglobulin receptor (alpha 2-MR). To analyze the generality of this mechanism, we studied the internalization of uPA by recombinant protease nexin-1 (rPN-1), an inhibitor of thrombin, uPA, and plasmin. 125I-uPA.rPN-1 complexes bound specifically to uPAR; internalization occurred efficiently, and its time course was essentially the same as for uPA.PAI-1. Internalization required binding to uPAR since it could be blocked by the anti-uPAR monoclonal antibodies, by the uPAR antagonist amino-terminal fragment of uPA, and by the removal of uPAR by the treatment of cells with phosphatidylinositol-specific phospholipase C. As for uPA.PAI-1, the internalization of uPA.rPN-1 also required alpha 2-MR, since it could be inhibited by the 39-kDa alpha 2-macroglobulin receptor/low density lipoprotein receptor-associated protein, a ligand for the alpha 2-MR. Finally, we show by ligand blot analysis that the uPA.rPN-1 complex, like uPA.PAI-1 but unlike free uPA, bound specifically to both uPAR and alpha 2-MR.
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PMID:Protease nexin-1-urokinase complexes are internalized and degraded through a mechanism that requires both urokinase receptor and alpha 2-macroglobulin receptor. 802 43

The amyloid beta-protein (A beta) and protease nexin-2/amyloid beta-protein precursor (PN-2/A beta PP) are major constituents of senile plaques and cerebrovascular deposits in individuals with Alzheimer's disease and related disorders. It has been suggested that the coagulation protease thrombin may process A beta PP in a manner leading to the formation of A beta. Here we investigated the effects of thrombin on the secretion and processing of PN-2/A beta PP and the production of A beta in a cellular system. Incubation of glioblastoma cells with thrombin (1-5 nM) resulted in the accumulation of abnormally processed, carboxyl-terminal-truncated forms of secreted PN-2/A beta PP (approximately 85 kDa) in the culture medium. Higher concentrations of thrombin (> 10 nM) also increased the levels of secreted PN-2/A beta PP in cultured untransfected glioblastoma cells and glioblastoma cells that were stably transfected to overproduce the 695 isoform of A beta PP. Increased secretion of PN-2/A beta PP required the proteolytic activity of thrombin, was induced by activation of the thrombin receptor by agonist peptides, and required activation of protein kinase C. Incubation of the untransfected and transfected glioblastoma cells with thrombin led to decreased levels of soluble A beta in the culture medium consistent with previously suggested mechanisms regarding the secretion of PN-2/A beta PP. Although the present studies suggest that thrombin does not directly contribute to A beta formation, its proteolysis of secreted PN-2/A beta PP may disrupt regions near the carboxyl terminus of the secreted proteins that account for their neuroprotective and cell adhesive properties.
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PMID:Thrombin receptor activation induces secretion and nonamyloidogenic processing of amyloid beta-protein precursor. 807 13

A 43 kd protein called glia-derived nexin or protease nexin-1 (GDN/PN-1) with both neurite-promoting and serine protease inhibitor activity is developmentally regulated during the differentiation of the nervous system. The synthesis of GDN/PN-1 remains high in structures such as the olfactory system where degeneration and regeneration take place throughout life. It is also up-regulated following injury both in the peripheral and the central nervous systems. Together with hirudin (a protease inhibitor from the leech), GDN/PN-1 is the most potent thrombin inhibitor known today. The surprising discovery of this potent thrombin inhibitor in the nervous system led to demonstration that mRNAs coding for prothrombin and the thrombin receptor are detected in neural tissue. Neuronal cells can cleave the inactive prothrombin into the active thrombin, which, in turn, specifically cleaves its own receptor to trigger a metabolic cascade causing sudden neurite retraction. Other macromolecules, such as vitronectin and thrombospondin, also found in the blood, can stimulate neurite outgrowth. Altogether, the data available today indicate that many molecules considered until now to be components of the hematopoietic system could perform distinct tasks and specific functions in the nervous system. Some of the experimental facts still required for demonstration of this hypothesis are discussed.
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PMID:Tinkering with certain blood components can engender distinct functions in the nervous system. 808 41


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