Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Prothrombin”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1990

Measurement of immunoreactive prothrombin, des-gamma-carboxy prothrombin, and vitamin K in human liver tissues: overproduction of immunoreactive prothrombin in hepatocellular carcinoma.

Des-gamma-carboxy prothrombin is an abnormal prothrombin which increases in the plasma of patients with hepatocellular carcinoma. To clarify the process of des-gamma-carboxy prothrombin synthesis, immunoreactive prothrombin, des-gamma-carboxy prothrombin, and vitamin K (phylloquinone and menaquinone) concentrations were determined in human liver tissue, including hepatocellular carcinoma. In the patients with elevated plasma des-gamma-carboxy prothrombin levels, both immunoreactive prothrombin and des-gamma-carboxy prothrombin significantly increased in hepatoma tissues compared with non-cancerous liver tissue. On the other hand, no significant difference was observed in the endogenous vitamin K (K1, MK-4, MK7) concentrations between hepatoma and noncancerous portions, in either the cases with or without increase of plasma des-gamma-carboxy prothrombin. These data strongly suggested that in the patients with an increase of plasma des-gamma-carboxy prothrombin, overproduction of prothrombin in hepatoma plays in important role in the synthesis of des-gamma-carboxy prothrombin.

Analysis of Variance↗
PubMed1976

Substitution of lanthanide ions for calcium ions in the activation of bovine prothrombin by activated factor X. High affinity metal-binding sites of prothrombin and the derivatives of prothrombin activation.

The substitution of lanthanide ions for Ca(II) in the Ca(II)-binding sites of prothrombin and the derivatives of prothrombin activation and in the metal-dependent conversion of prothrombin or prethrombin 1 to thrombin was studied at pH 6.8. Gd(III), Tb(III), La(III), Dy(III), Pr(III), Sm(III), and Ce(III) may be substituted for Ca(II) in the generation of thrombin from prothrombin or prethrombin 1 by activated factor X. The rates of thrombin generation in the presence of optimal concentrations of Gd(III) were about 25% for prothrombin and prethrombin 1 compared to the rate of thrombin generation with optimal concentrations of Ca(II). Maximal rates of thrombin generation were observed at 20 muM Gd(III) using prothrombin as substrate, compared to 10 muM Gd(III) when prethrombin 1 was employed. Using the steady state rate-dialysis method, the high affinity metal-binding sites of prothrombin and the products formed during prothrombin activation were characterized using 153Gd(III). Prothrombin has two high affinity binding sites for Gd(III) (Kd = 0.75 muM). Prethrombin 1 and prethrombin 2 each bind one Gd(III) tightly (Kd = 1.10 muM and 0.81 muM, respectively). Fragment 1, the phospholipid-binding portion of prothrombin, has two sites which bind Gd(III) tightly (Kd 0.16 muM). Fragment 2 has no high affinity metal-binding sites, but has intermediate affinity metal-binding sites (Kd greater than 1.6 muM). Thrombin has numerous high affinity binding sites (Kd less than 0.1 muM), suggesting that the conversion of prethrombin 2 to thrombin is associated with a significant change in tertiary structure. These results indicate that Gd(III) binds tightly to the metal-binding sites of these proteins and can substitute for Ca(II) in metal-dependent prothrombin activation. In the activation of prothrombin by activated factor X, these data suggest that Ca(II) is required for metal-dependent factor V and phospholipid binding and not as a cofactor in enzyme catalysis.

Animals↗
PubMed1983

Prothrombin Habana: a new dysfunctional molecule of human prothrombin associated with a true prothrombin deficiency.

A Cuban family with a new congenital dysprothrombinaemia is described. The propositus was a 5-year-old female who presented with umbilical bleeding after birth followed by easy bruising and bleeding tendency throughout her life. The main laboratory features of the defect included prolongation of prothrombin time and partial thromboplastin time. Prothrombin activity was less than 10% in several one- and two-stage systems. However, the staphylocoagulase-complexed prothrombin level and immunologic methods yielded levels of about 50%. The migration of the abnormal prothrombin was more anodic in single and bidimensional immunoelectrophoresis system and did not change by the addition of calcium. Family studies revealed that the father had approximately 50% prothrombin activity and antigen, whereas the mother had 45% prothrombin activity but about 100% prothrombin antigen. We suggest that the propositus is heterozygous for an abnormal prothrombin and heterozygous for true prothrombin deficiency.

Child, Preschool↗
PubMed1986

Prothrombin requires two sequential metal-dependent conformational transitions to bind phospholipid. Conformation-specific antibodies directed against the phospholipid-binding site on prothrombin.

Prothrombin is a gamma-carboxyglutamic acid-containing protein that binds to phospholipid vesicles in the presence of calcium ions after undergoing a metal ion-induced conformational transition. To integrate recent data into a scheme that is compatible with our knowledge of prothrombin-metal interaction, we have proposed a new model of prothrombin structure. In this model prothrombin undergoes two metal-dependent conformational transitions: PT----PT'----PT*. The first transition is not cation-specific, but the second transition is metal-selective for Ca(II), Sr(II), or Ba(II). Only the PT* conformer binds to phospholipid surfaces. To test this model, anti-prothrombin antibodies that only bind to prothrombin in the presence of Ca(II) but not Mg(II) (PT*-specific) were isolated, and termed anti-prothrombin X Ca(II)-specific. Half-maximal binding of antibody to prothrombin was observed at 0.1 mM CaCl2 or 1 mM SrCl2, but no binding was observed with Mg(II), Mn(II), or Ba(II). However, prothrombin in the presence of both Mg(II)/Ba(II) or Mn(II)/Ba(II) demonstrated significant interaction with the antibody. Prothrombin binding to phospholipid vesicles was inhibited by the anti-prothrombin X Ca(II)-specific antibody or its Fab fragment, but was not inhibited by anti-prothrombin X Mg(II) antibody or its Fab fragment directed at the PT' conformer. These results support this three-state model for prothrombin. The metal specificity characteristic of prothrombin-phospholipid interaction is a property required for the expression of the phospholipid-binding site in the binary prothrombin-metal complex.

Antibodies↗
PubMed1990

Randomized prospective trial comparing the native prothrombin antigen with the prothrombin time for monitoring oral anticoagulant therapy.

The dosage of the anticoagulant warfarin sodium is based upon the prolongation of the prothrombin time into an optimal therapeutic range. We have developed a new assay for the native prothrombin antigen that measures the fully gamma-carboxylated prothrombin using a radioimmunoassay. Based on preliminary data that indicated that the native prothrombin antigen predicted both bleeding and thrombotic complications more accurately than the prothrombin time in patients anticoagulated with warfarin sodium, we have performed a randomized prospective trial comparing the complication rate in warfarin-treated patients monitored with the native prothrombin antigen or the prothrombin time. Patients with indications for anticoagulation were randomized to be monitored by the native prothrombin antigen (therapeutic range, 12 to 24 micrograms/mL) or the prothrombin time index (therapeutic range, 1.5 to 2.0). Of the prothrombin time group (N = 80), seven (8.8%) had bleeding or thrombotic complications, with a complication rate of 9.5%/patient-year. In the native prothrombin antigen group (N = 76), one subject (1.3%) had a bleeding complication. The complication rate per patient-year was 1.5%. These results indicate an 85% reduction in the complication rate of the native prothrombin antigen group compared with the complication rate of the prothrombin time group. This difference is statistically significant by the Fisher exact test (P = .037) and by Kaplan Meier survival analysis (P = .040). This study suggests that the use of the native prothrombin antigen assay has the potential to decrease the complications associated with anticoagulation therapy with warfarin sodium.

Humans↗
PubMed1984

Monoclonal antibodies against human abnormal (des-gamma-carboxy)prothrombin specific for the calcium-free conformer of prothrombin.

A monoclonal antibody JO1 X 1 was prepared against human abnormal prothrombin using the hybridoma technique. The clone secreting this antibody was selected on the basis of the ability of this antibody to bind to abnormal prothrombin, but not to prothrombin, in the presence of calcium ions. The antibodies were purified by affinity chromatography in EDTA on columns of prothrombin-Sepharose. Bound antibodies were eluted with 15 mM CaCl2. The kinetics of dissociation of antibody from the antibody-prothrombin complex with the addition of calcium ions fit a first-order kinetic model. Increasing CaCl2 concentration increased the rate of antibody-prothrombin dissociation. Ca(II) and Mn(II) inhibited antibody-prothrombin interaction; half-maximal binding was observed at 0.9 and 4 mM, respectively. Mg(II) had little effect on antibody-antigen interaction. The JO1 X 1 antibody bound fragment 1, fragment (1-39), abnormal prothrombin, and prothrombin equivalently in the presence of EDTA, but did not bind to des(1-44)prothrombin in the presence of EDTA or prothrombin in the presence of CaCl2. These results indicate that the monoclonal antibody JO1 X 1 is conformation specific for the calcium-free conformer of prothrombin and directed against an antigenic determinant near the NH2 terminus of prothrombin expressed in the 1-39 region of the protein. This analysis provides confirmation of the presence of a metal-free conformer of prothrombin.

Animals↗
PubMed1992

Relationship between total prothrombin, native prothrombin and the International Normalized Ratio (INR).

Plasma levels of total prothrombin and fully-carboxylated (native) prothrombin were compared with results of prothrombin time (PT) assays for patients undergoing oral anticoagulant therapy. Mean concentrations of total and native prothrombin in non-anticoagulated patients were 119 +/- 13 micrograms/ml and 118 +/- 22 micrograms/ml, respectively. In anticoagulated patients, INR values ranged as high as 9, and levels of total prothrombin and native prothrombin decreased with increasing INR to minimum values of 40 micrograms/ml and 5 micrograms/ml, respectively. Des-carboxy-prothrombin increased with INR, to a maximum of 60 micrograms/ml. The strongest correlation was observed between native prothrombin and the reciprocal of the INR (1/INR) (r = 0.89, slope = 122 micrograms/ml, n = 200). These results indicated that native prothrombin varied over a wider range and was more closely related to INR values than either total or des-carboxy-prothrombin. Levels of native prothrombin were decreased 2-fold from normal levels at INR = 2, indicating that the native prothrombin antigen assay may be a sensitive method for monitoring low-dose oral anticoagulant therapy. The inverse relationship between concentration of native prothrombin and INR may help in identification of appropriate therapeutic ranges for oral anticoagulant therapy.

Administration, Oral↗
PubMed1984

Comparison of the native prothrombin antigen and the prothrombin time for monitoring oral anticoagulant therapy.

We have measured the fully carboxylated (native) prothrombin antigen and the undercarboxylated (abnormal) prothrombin antigen in patients treated with sodium warfarin using specific immunoassays to evaluate a new approach for monitoring oral anticoagulant therapy. Plasma and serum samples (391) were assayed for the prothrombin time, native prothrombin antigen, and abnormal prothrombin antigen. The results were correlated with the presence of bleeding or thromboembolic complications at the time of phlebotomy. The native prothrombin antigen correlated with the occurrence of complications in 95% of samples. Of 13 samples from patients with bleeding complications, 13/13 (100%) had a native prothrombin of 12 micrograms/mL or lower. Of seven samples from patients with thromboembolic complications, 6/7 (86%) had a native prothrombin of 24 micrograms/mL or greater. By comparison, a prothrombin time index of 1.5 to 2.5, 1.5 to 2.2, 1.5 to 2.0, or 1.3 to 1.8 identified 6/20 (30%), 9/20 (45%), 11/20 (55%), or 12/20 (60%) patients at risk, respectively. Although the prothrombin time index did correlate with the presence of bleeding complications, the native prothrombin antigen correlated closely with the presence of bleeding and thromboembolic complications. According to these results, the native prothrombin antigen, maintained in a range of 12 to 24 micrograms/mL by regular adjustment of the warfarin dosage, may be associated with a reduced risk of complications due to excessive or insufficient warfarin therapy. On the basis of these preliminary data, we recommend that the native prothrombin antigen be considered to monitor warfarin therapy.

Administration, Oral↗
PubMed1985

The effects of bovine prothrombin fragment 1 and fragment 1.2 on prothrombin activation.

In this paper we describe the effects of the activation peptides prothrombin fragment 1 and fragment 1.2 on factor Xa-catalyzed prothrombin activation. Prothrombin activation in free solution by either factor Xa or factor Xa together with factor Va is unaffected by the activation fragments. When negatively charged phospholipids are present we observed considerable inhibition of prothrombin activation by both fragment 1 and fragment 1.2. For the activation of 0.25 microM prothrombin by factor Xa in the presence of 50 microM phospholipid (phosphatidylserine/phosphatidylcholine, 25/75; mol/mol) and 5 mM CaCl2 50% inhibition was obtained at 0.28 microM fragment 1 or fragment 1.2. Much higher fragment concentrations were required for 50% inhibition of a prothrombinase complex consisting of factor Xa, factor Va, Ca2+ and phospholipid. This shows that factor Va protects prothrombin activation against inhibition by its own activation peptides. Less inhibition by activation fragments was also observed at higher phospholipid and prothrombin concentrations or when the mole fraction phosphatidylserine in the phospholipid vesicles was decreased. The effects of fragment 1 and fragment 1.2 on prothrombin activation were identical throughout all experiments, indicating that the inhibition is due to the gamma-carboxyglutamic acid containing region of the activation peptides. Our observations suggest that the activation fragments inhibit prothrombin activation by competing with prothrombin and factor Xa for binding sites at the phospholipid surface. In such a model factor Va will protect against the inhibition since it is known to promote the assembly of the prothrombinase complex through interactions with factor Xa and prothrombin that are independent of the gla-residues. The kinetic properties of fragment inhibition also suggest that in vivo prothrombin activation will not be affected by the generation of activation peptides.

Animals↗
PubMed1991

Prothrombin Himi; an abnormal prothrombin characterized by a defective thrombin activity.

An abnormal prothrombin has been detected in a 26-year-old female, who had no history of excessive bleeding. Prothrombin activity was approximately 10% when measured using either the classical one-stage assay or the assay with Echis carinatus venom, whereas prothrombin antigen level was normal. In keeping with current nomenclature practices, the abnormal prothrombin was designated "Prothrombin Himi". The electrophoretic behavior and calcium binding properties of Prothrombin Himi did not differ significantly from normal. Prothrombin Himi was isolated by chromatography on Q-Sepharose. Electrophoretic migration of the purified abnormal prothrombin on SDS-PAGE was normal. Upon prothrombin activation by Echis carinatus venom, the clotting activity produced from Prothrombin Himi was only 37% of the normal level after 90 minutes of the activation time, where as the amidolytic activity was almost the same as normal. The cleavage patterns of Prothrombin Himi by factor Xa or Echis carinatus venom investigated by SDS-PAGE, were found to be normal. These results indicate that Prothrombin Himi was characterized by a defective thrombin enzymatic activity.

Adult↗
PubMed1995

Differences in the interactions of lupus anticoagulant IgG with human prothrombin and bovine prothrombin.

Lupus anticoagulant (LA) IgGs have been reported to inhibit more effectively and consistently the Xa/Va/phospholipid complex-catalyzed activation of human prothrombin that the Xa/Va/phospholipid complex-catalyzed activation of bovine prothrombin. This led us to carry out studies to determine whether the ability to inhibit the activation of prothrombin of LA IgGs, separated from the plasma of 15 patients by protein A affinity chromatography, could be related to the ability of the LA IgGs to bind to prothrombin under various experimental conditions. Of 14 LA IgG preparations tested all prolonged to a variable but substantial extent the dilute Russell's viper venom time (dRVVT) of human plasma but only minimally prolonged the dRVVT of bovine plasma. In a purified prothrombin activation system with a rate limiting concentration of phospholipid, all 15 LA IgG preparations inhibited the activation of human prothrombin with the majority showing > 50% of inhibition. In contrast, only one LA IgG markedly inhibited (> 50%) the activation of bovine prothrombin and five others moderately inhibited (25-40%) the activation of bovine prothrombin. Nevertheless, the majority of LA IgG preparations bound to immobilized bovine prothrombin on a Western blot and also to immobilized bovine prothrombin on a microtiter well. In an ELISA in which phosphatidylserine (PS) was immobilized on microtiter wells, bovine prothrombin supported the binding of 10 of 15 LA IgG preparations to PS. However, the extent of binding was lower than that observed with human prothrombin.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Prothrombin Salakta: an abnormal prothrombin characterized by a defect in the active site of thrombin.

An abnormal prothrombin has been detected in a 17 yr-old female originating from Tunisia. There was no history of excessive bleeding. Prothrombin time and activated partial thromboplastin time were moderately prolonged. Prothrombin activity was 15-18% when measured using either the classical one-stage and two-stage assays, or assays with Echis carinatus venom or staphylocoagulase, whereas prothrombin antigen was 100%. In keeping with current nomenclature practices, the abnormal molecule has been designated prothrombin Salakta. The electrophoretic behaviour and calcium binding properties of the abnormal prothrombin did not differ significantly from normal, as assessed by crossed immunoelectrophoresis. Prothrombin Salakta was isolated by chromatography on DEAE-Sephadex and Dextran sulphate sepharose. Electrophoretic migration of purified prothrombin Salakta on SDS polyacrylamide gels or alkaline disc gels was normal. Upon activation by either bovine factor Xa or Echis carinatus venom, thrombin activity produced by prothrombin Salakta was only 15% of normal, even when the incubation period was prolonged for 24 hours. The pattern of factor Xa-catalyzed proteolysis of prothrombin Salakta, investigated by SDS polyacrylamide gel electrophoresis, was found to be normal. These results indicated that prothrombin Salakta was characterized by a defective thrombin enzymatic activity. Thrombin Salakta was therefore isolated by heparin-sepharose chromatography. Affinity for heparin and molecular weight of thrombin Salakta were found to be normal. Biological activity of thrombin Salakta, determined by clotting assay, was 535 u/mg versus 3 200 u/mg for normal thrombin. Amidolytic activity of thrombin Salakta parallelled its clotting activity, suggesting that the defect resides either in the catalytic site or in the residues adjacent to the catalytic site and implicated as contact residues, rather than in the fibrinogen recognition site.

Adolescent↗
PubMed1991

Modulation of human prothrombin activation on phospholipid vesicles and platelets using monoclonal antibodies to prothrombin fragment 2.

Prothrombin contains two kringle domains that are removed during activation to the blood clotting enzyme alpha-thrombin. By analogy with other kringle-containing proteins the prothrombin kringles may play a role in the protein-protein interactions necessary for prothrombin activation. Four monoclonal antibodies to prothrombin kringle 2 have been produced against human prothrombin, and a fifth monoclonal antibody was produced against a synthetic peptide consisting of amino acid residues 216-231 of kringle 2. Each antibody was tested for its ability to block prothrombin activation by factor Xa. In the presence of phosphatidylcholine/phosphatidylserine vesicles and factor Va, two of the antibodies, alpha HII-3 and alpha HII-4, inhibited prothrombin activation at a 90 and 50% level, respectively. Two other monoclonal antibodies (alpha HII-6 and alpha HII-7) and the antipeptide antibody (alpha HII-5) had no effect on prothrombin activation. When factor Xa was the catalyst alone, antibody alpha HII-3 lost the ability to inhibit prothrombin activation whereas antibody alpha HII-4 again partially inhibited the reaction. When human platelets were the reaction surface, the patterns of inhibition by the anti-fragment 2 antibodies were identical to that observed with phospholipid vesicles. These data suggest a role for prothrombin fragment 2 in activation, possibly by mediating the interaction of substrate prothrombin with factor Xa or factor Va on the phospholipid surface.

Amino Acid Sequence↗
PubMed1986

Prothrombin fragment 1 X 2 X 3, a major product of prothrombin activation in human plasma.

The conversion of the blood coagulation zymogen prothrombin to thrombin is associated with the production of several cleavage intermediates and products. In contrast to earlier studies of prothrombin cleavage in chemically defined systems, the current investigation examines the fragmentation of human prothrombin in normal plasma. Radiolabeled prothrombin was added to platelet-poor relipidated normal human plasma, and clotting was initiated with the addition of Ca(II) and kaolin. Analysis of the radiolabeled prothrombin cleavage products by polyacrylamide gel electrophoresis in the presence of dodecyl sulfate and beta-mercaptoethanol identified a heretofore unobserved product of prothrombin activation with an apparent molecular weight of 45,000. This product was identified as fragment 1 X 2 X 3, the NH2-terminal 286 amino acids of prothrombin. The product was isolated from a prothrombin digest by immunoaffinity chromatography using anti-prothrombin:Ca(II) antibodies and by preparative gel electrophoresis. Its amino-terminal sequence is identical to that of prothrombin. Digestion of this product with either Factor Xa or thrombin yields, at a minimum, fragment 1 X 2 and fragment 1. Amino-terminal sequence analysis of the products obtained by digestion with Factor Xa of the unknown activation product indicated 3 amino acid residues at each cycle consistent with the presence of fragment 1, fragment 2, and fragment 3. To unambiguously identify the COOH-terminal amino acid sequence of the product, its factor Xa digestion products were separated by reverse-phase high performance liquid chromatography. Edman degradation of one peptide revealed the complete sequence of fragment 3. On this basis, we identify the Mr 45,000 polypeptide as fragment 1 X 2 X 3 and indicate that it is a prominent product of prothrombin conversion to thrombin when activation occurs in plasma.

Amino Acid Sequence↗
PubMed1987

Homo- and heterodimer formation with prothrombin and prothrombin fragment 1 in the presence of calcium ions.

The purpose of the current study is to present further evidence for prothrombin self-association as assessed by chemical crosslinking. When the self-association (evaluated by covalent crosslinking with dithiobis(succinimidylpropionate) of prothrombin or fragment 1 was evaluated at the same molar concentration of protein, similar rates of dimer formation were observed for either protein. When prothrombin and fragment 1 were incubated together with the crosslinking reagent and calcium ions, a heterodimer consisting of prothrombin and fragment 1 was observed in addition to prothrombin dimer and fragment 1 dimer. Similar experiments with prethrombin 1 showed neither significant self-association nor effect on prothrombin self-association. Comparison of the formation of prothrombin fragment 1 heterodimer formation with the effect of fragment 1 on prothrombin activation by factor Xa suggests that the anticoagulant activity of fragment 1 is not solely a result of the formation of a heterodimer between prothrombin and fragment 1.

Animals↗
PubMed1991

Fourier transform infrared spectroscopic study of Ca2+ and membrane-induced secondary structural changes in bovine prothrombin and prothrombin fragment 1.

Fourier transform infrared (FTIR) spectroscopy was used to monitor secondary structural changes associated with binding of bovine prothrombin and prothrombin fragment 1 to acidic lipid membranes. Prothrombin and prothrombin fragment 1 were examined under four different conditions: in the presence of (a) Na2EDTA, (b) 5 mM CaCl2, and in the presence of CaCl2 plus membranes containing 1-palmitoyl-2-oleoyl-3-sn-phosphatidylcholine (POPC) in combination with either (c) bovine brain phosphatidyl-serine (bovPS) or (d) 1,2-dioleoyl-phosphatidylglycerol (DOPG). The widely reported Ca(2+)-induced conformational change in bovine prothrombin fragment 1 was properly detected by our procedures, although Ca(2+)-induced changes in whole prothrombin spectra were too small to be reliably interpreted. Binding of prothrombin in the presence of Ca2+ to procoagulant POPC/bovPS small unilamellar vesicles produced an increase in ordered secondary structures (2% and 3% increases in alpha-helix and beta-sheet, respectively) and a decrease of random structure (5%) as revealed by spectral analysis on both the original and Fourier-self-deconvolved data and by difference spectroscopy with the undeconvolved spectra. Binding to POPC/DOPG membranes, which are less active as procoagulant membranes, produced no detectable changes in secondary structure. In addition, no change in prothrombin fragment 1 secondary structure was detectable upon binding to either POPC/bovPS or POPC/DOPG membranes. This indicates that a membrane-induced conformational change occurs in prothrombin in the nonmembrane-binding portion of the molecule, part of which is activated to form thrombin, rather than in the membrane-binding fragment 1 region. The possible significance of this conformational change is discussed in terms of differences between the procoagulant activities of different acidic lipid membranes.

Animals↗