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Nihon Rinsho. Japanese Journal of... Sep 1999
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The Journal of Biological Chemistry Aug 2020The receptor for advanced glycation end products (RAGE) plays a key role in mammal physiology and in the etiology and progression of inflammatory and oxidative...
The receptor for advanced glycation end products (RAGE) plays a key role in mammal physiology and in the etiology and progression of inflammatory and oxidative stress-based diseases. In adults, RAGE expression is normally high only in the lung where the protein concentrates in the basal membrane of alveolar Type I epithelial cells. In diseases, RAGE levels increase in the affected tissues and sustain chronic inflammation. RAGE exists as a membrane glycoprotein with an ectodomain, a transmembrane helix, and a short carboxyl-terminal tail, or as a soluble ectodomain that acts as a decoy receptor (sRAGE). VC1 domain is responsible for binding to the majority of RAGE ligands including advanced glycation end products (AGEs), S100 proteins, and HMGB1. To ascertain whether other ligands exist, we analyzed by MS the material pulled down by VC1 from human plasma. Twenty of 295 identified proteins were selected and associated to coagulation and complement processes and to extracellular matrix. Four of them contained a γ-carboxyl glutamic acid (Gla) domain, a calcium-binding module, and prothrombin (PT) was the most abundant. Using MicroScale thermophoresis, we quantified the interaction of PT with VC1 and sRAGE in the absence or presence of calcium that acted as a competitor. PT devoid of the Gla domain (PT des-Gla) did not bind to sRAGE, providing further evidence that the Gla domain is critical for the interaction. Finally, the presence of VC1 delayed plasma clotting in a dose-dependent manner. We propose that RAGE is involved in modulating blood coagulation presumably in conditions of lung injury.
Topics: Blood Coagulation; Humans; Lung Injury; Protein Binding; Protein Domains; Prothrombin; Receptor for Advanced Glycation End Products
PubMed: 32665403
DOI: 10.1074/jbc.RA120.013692 -
Haemostasis 2001We have recently determined the complete amino acid sequence of trocarin, a group D prothrombin activator from the venom of Tropidechis carinatus (Australian... (Review)
Review
We have recently determined the complete amino acid sequence of trocarin, a group D prothrombin activator from the venom of Tropidechis carinatus (Australian rough-scaled snake). This proteinase is both functionally and structurally similar to mammalian blood coagulation factor Xa. It shows approximately 70% homology and possesses the characteristic Gla domain, two EGF domains and serine proteinase domain. To examine structure-function relationships, we generated a molecular model of trocarin based on a human factor Xa des-Gla crystal structure (1xka) as template. Based on known sites of interaction between mammalian factor Xa, factor Va and prothrombin, structure-function relationships of trocarin were explored. Unlike factor Xa, trocarin is glycosylated and has a large carbohydrate moiety at the entrance to its active site pocket. This might contribute to differences observed in the kinetics of hydrolysis of synthetic substrates by trocarin as compared to human factor Xa. A Ca(2+)-binding loop present in the heavy chain of factor Xa also seems to be lost in trocarin. In addition to its role in hemostasis, factor Xa shows other biological effects, including inflammation via its interaction with effector protease receptor-1 (EPR-1). Interestingly, the EPR-1 recognition site is distinctly different in trocarin, the functional consequences of which are being investigated.
Topics: Animals; Binding Sites; Coagulants; Factor Xa; Humans; Protein Processing, Post-Translational; Prothrombin; Sequence Homology, Nucleic Acid; Snake Venoms
PubMed: 11910190
DOI: 10.1159/000048068 -
Medecine Et Hygiene May 1950
Topics: Hemostatics; Prothrombin
PubMed: 15416281
DOI: No ID Found -
Scientific Reports Feb 2018The clotting factor prothrombin exists in equilibrium between closed and open conformations, but the physiological role of these forms remains unclear. As for other...
The clotting factor prothrombin exists in equilibrium between closed and open conformations, but the physiological role of these forms remains unclear. As for other allosteric proteins, elucidation of the linkage between molecular transitions and function is facilitated by reagents stabilized in each of the alternative conformations. The open form of prothrombin has been characterized structurally, but little is known about the architecture of the closed form that predominates in solution under physiological conditions. Using X-ray crystallography and single-molecule FRET, we characterize a prothrombin construct locked in the closed conformation through an engineered disulfide bond. The construct: (i) provides structural validation of the intramolecular collapse of kringle-1 onto the protease domain reported recently; (ii) documents the critical role of the linker connecting kringle-1 to kringle-2 in stabilizing the closed form; and (iii) reveals novel mechanisms to shift the equilibrium toward the open conformation. Together with functional studies, our findings define the role of closed and open conformations in the conversion of prothrombin to thrombin and establish a molecular framework for prothrombin activation that rationalizes existing phenotypes associated with prothrombin mutations and points to new strategies for therapeutic intervention.
Topics: Amino Acid Sequence; Enzyme Activation; Enzyme Precursors; Enzyme Stability; Humans; Kinetics; Models, Molecular; Mutation; Protein Conformation; Protein Engineering; Prothrombin; Structure-Activity Relationship; Thrombin
PubMed: 29440720
DOI: 10.1038/s41598-018-21304-1 -
La Revue de Medecine Interne Sep 2022Antiphospholipid syndrome (APS) is an autoimmune disease and one of the most common causes of acquired thrombophilia. It is characterised by the occurrence of thrombotic...
Antiphospholipid syndrome (APS) is an autoimmune disease and one of the most common causes of acquired thrombophilia. It is characterised by the occurrence of thrombotic or obstetric events associated with the presence of persistent antiphospholipid antibodies. The diagnosis can be challenging, particularly because some biological tests can be disturbed by anticoagulant treatment or inflammation. In the recent years, new antiphospholipid antibodies, including anti-phosphatidylserine/prothrombin antibodies (anti-PS/PT), have emerged but their clinical significance and causality remain uncertain. Biologically, several studies have found a strong correlation between the presence of lupus anticoagulant (LA) and anti-PS/PT antibodies. Clinically, the presence of anti-PS/PT antibodies is associated with an increased risk of thrombosis and obstetric complications. There is also an association with thrombocytopenia, suggesting that the presence of anti-PS/PT antibodies may be associated with more severe clinical APS. Among seronegative APS patients, 6-17% of patients are positive for anti-PS/PT antibodies. This might influence the therapeutic management of patients. This article aims to provide an update on contribution of anti-PS/PT antibodies detection for the diagnosis and management of APS.
Topics: Antibodies, Antiphospholipid; Antiphospholipid Syndrome; Female; Humans; Lupus Coagulation Inhibitor; Phosphatidylserines; Pregnancy; Prothrombin; Thrombosis
PubMed: 35752484
DOI: 10.1016/j.revmed.2022.04.031 -
Thrombosis Research Nov 2017Antithrombin resistance (ATR) prothrombinemia is an inherited thrombophilic disorder caused by missense mutations in prothrombin gene (F2) at Arg596 of the...
INTRODUCTION
Antithrombin resistance (ATR) prothrombinemia is an inherited thrombophilic disorder caused by missense mutations in prothrombin gene (F2) at Arg596 of the sodium-binding region. Previously, prothrombin mutants Yukuhashi (Arg596Leu), Belgrade (Arg596Gln), and Padua 2 (Arg596Trp) were reported as ATR-prothrombins possessing a risk of familial venous thrombosis. To identify additional F2 mutations causing the ATR-phenotype, we investigated the coagulant properties of recombinant prothrombins mutated at amino acid residues within the sodium-binding region by single nucleotide substitutions (Thr540, Arg541, Glu592, and Lys599).
MATERIALS AND METHODS
We constructed expression vectors of prothrombin mutants, established stably transfected HEK293 cells, and isolated the recombinant prothrombin proteins. We evaluated procoagulant activity and ATR-phenotypes of those mutants in reconstituted plasma by mixing with prothrombin deficient plasma.
RESULTS
The secreted quantity of all prothrombin mutants was the same as that of the wild-type prothrombin. Procoagulant activity of each mutant varied from 1.7% to 79.5% in a one-stage clotting assay and from 2.0% to 104.5% in a two-stage chromogenic assay. Most prothrombin mutants tested presented with a severe ATR-phenotype. To estimate the thrombosis risk of these mutations, we determined the residual clotting activity (RCA) after 30min inactivation with antithrombin. RCA scores, normalized to the wild-type, revealed that prothrombin mutants Lys599Arg (5.35) and Glu592Gln (4.71) had high scores, which were comparable with prothrombins Yukuhashi (4.36) and Belgrade (5.19).
CONCLUSIONS
Mutation of prothrombin at the sodium-binding site caused ATR-phenotypes. Of those tested, Lys599Arg and Glu592Gln may possess a thrombosis risk as large as the known pathogenic prothrombins Yukuhashi and Belgrade.
Topics: Antithrombins; Drug Resistance, Neoplasm; Humans; Mutation; Prothrombin
PubMed: 28961453
DOI: 10.1016/j.thromres.2017.09.020 -
The New England Journal of Medicine Feb 1974
Topics: Adult; Blood Coagulation Factors; Blood Coagulation Tests; Hemophilia B; Humans; Infusions, Parenteral; Male; Middle Aged; Postoperative Complications; Prothrombin; Thrombosis
PubMed: 4810120
DOI: 10.1056/NEJM197402142900718 -
The Journal of Biological Chemistry Aug 2016The coagulation factor prothrombin has a complex spatial organization of its modular assembly that comprises the N-terminal Gla domain, kringle-1, kringle-2, and the...
The coagulation factor prothrombin has a complex spatial organization of its modular assembly that comprises the N-terminal Gla domain, kringle-1, kringle-2, and the C-terminal protease domain connected by three intervening linkers. Here we use single molecule Förster resonance energy transfer to access the conformational landscape of prothrombin in solution and uncover structural features of functional significance that extend recent x-ray crystallographic analysis. Prothrombin exists in equilibrium between two alternative conformations, open and closed. The closed conformation predominates (70%) and features an unanticipated intramolecular collapse of Tyr(93) in kringle-1 onto Trp(547) in the protease domain that obliterates access to the active site and protects the zymogen from autoproteolytic conversion to thrombin. The open conformation (30%) is more susceptible to chymotrypsin digestion and autoactivation, and features a shape consistent with recent x-ray crystal structures. Small angle x-ray scattering measurements of prothrombin wild type stabilized 70% in the closed conformation and of the mutant Y93A stabilized 80% in the open conformation directly document two envelopes that differ 50 Å in length. These findings reveal important new details on the conformational plasticity of prothrombin in solution and the drastic structural difference between its alternative conformations. Prothrombin uses the intramolecular collapse of kringle-1 onto the active site in the closed form to prevent autoactivation. The open-closed equilibrium also defines a new structural framework for the mechanism of activation of prothrombin by prothrombinase.
Topics: Amino Acid Substitution; Humans; Mutation, Missense; Protein Domains; Prothrombin; X-Ray Diffraction
PubMed: 27435675
DOI: 10.1074/jbc.M116.738310 -
Vitamins and Hormones 1974
Review
Topics: Amino Acids; Animals; Cycloheximide; Enzyme Precursors; Liver; Microsomes, Liver; Models, Chemical; Prothrombin; Vitamin K; Vitamin K Deficiency; Warfarin
PubMed: 4617405
DOI: 10.1016/s0083-6729(08)60023-0