scholarly journals The Effect of Fibrinopeptide B Release on Temperature Dependent Fibrin Association

1977 ◽  
Author(s):  
W. Edgar ◽  
C.R.M. Prentice
Keyword(s):  
Affinity Chromatography ◽  
Clot Formation ◽  
Effect Of Temperature ◽  
Fibrinopeptide A ◽  
Temperature Dependent ◽  
Fibrin Polymerization ◽  
Soluble Fibrin ◽  
Agkistrodon Contortrix ◽  
The Stability ◽  
Sepharose 4B

The effect of temperature on soluble fibrin complexes was studied using Biogel filtration and chromatography on fibrinogen-sepharose 4B at 20°C and 37°C. Complexes were formed by plasmin digestion of non-crosslinked fibrin produced by thrombin, ancrod, thrombin in 2M urea, or ancrod plus Agkistrodon contortrix venom. Thrombin removes fibrinopeptides A and B, ancrod removes fibrinopeptide A, while A. contortrix enzyme removes first fibrinopeptide B, followed by A. Complexes containing neither fibrinopeptide A or B, formed by digestion of fibrin produced by thrombin, or by ancrod plus Agkistrodon contortrix, were stable at 37°C. In contrast, complexes which retained fibrinopeptide B, formed from fibrin produced by ancrod or by thrombin in 2M urea, were unstable at 37°C. Fibrin polymerization was necessary for the stability of fibrin complexes. Complexes from plasmin digests of fibrin produced by ancrod plus A. contortrix enzyme in 2M urea, where no clot formation occurred, were unstable at 37°C. Using affinity chromatography, plasmin digests of thrombin-fibrin bound to fibrinogen-sepharose at 37°C, whereas those from ancrod-fibrin did not. A second set of polymerization sites in fibrinogen are proposed, distinct from the N-DSK and carboxyterminal sites. These are activated by removal of fibrinopeptide B and require clot formation.

Quantitative Assessment of Soluble Fibrin in Plasma by Affinity Chromatography – A Comparative Study with desAA-Fibrin, desAABB-Fibrin and Fibrinogen

1985 ◽  
Vol 54 (02) ◽  
pp. 533-538 ◽  
Author(s):  
Wilfried Thiel ◽  
Ulrich Delvos ◽  
Gert Müller-Berghaus
Keyword(s):  
Affinity Chromatography ◽  
Calcium Ions ◽  
Quantitative Assessment ◽  
Fluid Phase ◽  
Soluble Fibrin ◽  
Binding Characteristics ◽  
Different Temperatures ◽  
Immobilized Proteins ◽  
Sepharose 4B

SummaryA quantitative determination of soluble fibrin in plasma was carried out by affinity chromatography. For this purpose, desAA-fibrin and fibrinogen immobilized on Sepharose 4B were used at the stationary side whereas batroxobin-induced 125I-desAA-fibrin or thrombin-induced 125I-desAABB-fibrin mixed with plasma containing 131I-fibrinogen represented the fluid phase. The binding characteristics of these mixtures to the immobilized proteins were compared at 20° C and 37° C. Complete binding of both types of fibrin to the immobilized desAA-fibrin was always seen at 20° C as well as at 37° C. However, binding of soluble fibrin was accompanied by substantial binding of fibrinogen that was more pronounced at 20° C. Striking differences depending on the temperature at which the affinity chromatography was carried out, were documented for the fibrinogen-fibrin interaction. At 20° C more than 90% of the applied desAA-fibrin was bound to the immobilized fibrinogen whereas at 37° C only a mean of 17% were retained at the fibrinogen-Sepharose column. An opposite finding with regard to the tested temperature was made with the desAABB-fibrin. Nearly complete binding to insolubilized fibrinogen was found at 37° C (95%) but only 58% of the desAABB-fibrin were bound at 20° C. The binding patterns did not change when the experiments were performed in the presence of calcium ions. The opposite behaviour of the two types of soluble fibrin to immobilized fibrinogen at the different temperatures, together with the substantial binding of fibrinogen in the presence of soluble fibrin to insolubilized fibrin in every setting tested, devaluates affinity chromatography as a tool in the quantitative assessment of soluble fibrin in patients’ plasma.

Improved purification of human lactate dehydrogenase isoenzyme-3 and studies with its fluorescein isothiocyanate and biotin conjugates

1990 ◽  
Vol 36 (1) ◽  
pp. 59-64
Author(s):  
R N Weijers ◽  
R de Bruijn ◽  
J Mulder ◽  
H Kruijswijk
Keyword(s):  
Amino Acid ◽  
Lactate Dehydrogenase ◽  
Affinity Chromatography ◽  
B Lymphocytes ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Fluorescein Isothiocyanate ◽  
Molar Ratio ◽  
The Stability ◽  
Sepharose 4B

Abstract Lactate dehydrogenase (L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) isoenzyme-3 (LD-3) has been isolated in milligram quantities from human erythrocytes. Using an improved procedure--which involves complete hemolysis of the erythrocytes, diethylaminoethyl (DEAE)-Sephacel column chromatography, and 5'-AMP-Sepharose 4B affinity chromatography--we obtained 23,000-fold purified isoenzyme from the crude hemolysate (overall yield about 90%). The final product was homogeneous on polyacrylamide disc gel electrophoresis and had a specific activity of about 435 kU/g. Its amino acid composition is presented. With the eventual aim to make visible and isolate IgA kappa antibody-secreting B lymphocytes, we developed reproducible methods for preparing fluorescein isothiocyanate isomer-1-conjugated LD-3 with a fluorescein/LD-3 molar ratio between 1.3 and 3.3, and biotinylated LD-3 with a biotin/LD-3 molar ratio between 1.3 and 2.5. In evaluating the stability of these two conjugates, we determined that they still can react with IgA kappa to form the IgA kappa (LD-3)2 complex.

The use of Spheron as a matrix for affinity chromatography of NAD-dependent dehydrogenases

1986 ◽  
Vol 51 (7) ◽  
pp. 1542-1549
Author(s):  
Jan Kovář ◽  
Alena Škodová ◽  
Jaroslav Turánek
Keyword(s):  
Lactate Dehydrogenase ◽  
Alcohol Dehydrogenase ◽  
Affinity Chromatography ◽  
Dehydrogenase Activity ◽  
Nitrous Acid ◽  
Separation Efficiency ◽  
Binding Capacity ◽  
The Stability ◽  
Sepharose 4B

The paper compares several methods of coupling common ligands of dehydrogenases, viz. N6-[(6-aminohexyl)carbamoylmethyl]-AMP and N6-[(6-aminohexyl)carbamoylmethyl]-NAD, to a hydrophilic macroporous glycolmethacrylate gel, Spheron. The affinants coupled best to a CNBr-activated gel and to a gel with hydrazine groups (after activation with nitrous acid). The affinity properties of gels based on Spheron and on Sepharose 4B were similar ( the stability and separation efficiency were almost identical, the binding capacity and the recovery of dehydrogenase activity were somewhat better with the Sepharose). The materials based on Spheron were used in several separation experiments, viz. separation of lactate dehydrogenase form albumin, separation of lactate dehydrogenase from alcohol dehydrogenase under different conditions and separation of isoenzymes of lactate dehydrogenase. Spheron 300 with a coupled affinant was also employed in an attempt to purify a crude alcohol dehydrogenase.

scholarly journals On the Modulation of Oscillation in Thermohaline Convection Problems Using Temperature Dependent Viscosity

2015 ◽  
Vol 45 (1) ◽  
pp. 39-52
Author(s):  
Joginder Singh Dhiman ◽  
Vijay Kumar
Keyword(s):  
Growth Rate ◽  
Linear Stability Theory ◽  
Effect Of Temperature ◽  
Sufficient Condition ◽  
Temperature Dependent ◽  
Temperature Dependent Viscosity ◽  
Thermohaline Convection ◽  
The Stability ◽  
Dependent Viscosity ◽  
Complex Growth Rate

Abstract The present paper mathematically investigates the effect of temperature dependent viscosity on the onset of instability in thermohaline convection problems of Veronis and Stern type configurations, using linear stability theory. A sufficient condition for the stability of oscillatory modes for thermohaline configuration is derived. When the compliment of this sufficient condition is true, the oscillatory motions of neutral or growing amplitude may exist, and hence the bounds for the complex growth rate of these neutral or unstable modes are derived, when viscosity of the fluid is an arbitrary function of temperature. Some general conclusions for the cases of linear and exponential variations of viscosity are worked out. The present analysis thus shows that the oscillations in thermohaline convection problems can be modulated or arrested by considering the temperature dependent viscosity of the fluid.

Sequential Formation Of Des-A Fibrin And Des-Ab Fibrin And Binding Of Fibrinogen To Des-Ab Fibrin

1981 ◽  
Author(s):  
G Müller-Berghaus ◽  
J C Bernhard ◽  
H Hofmann ◽  
I Mahn ◽  
F R Ostheimer ◽  
...  
Keyword(s):  
Affinity Chromatography ◽  
Weak Interaction ◽  
Gel Filtration ◽  
Clot Formation ◽  
Soluble Fibrin ◽  
Low Concentration ◽  
Thrombin Cleavage ◽  
Physico Chemical ◽  
Sequential Formation

Thrombin cleaves the fibrinopeptides A and B from fibrinogen in a consecutive manner, producing first des-A fibrin and consequently, des-AB fibrin. Physico-chemical and in-vivo properties of both types of fibrin were comparatively studied by gel filtration, affinity chromatography and in-vivo behavior. At 37°C and in the presence of fibrinogen and Ca++, des-A fibrin formed soluble fibrin-fibrin dimers. Des-AB fibrin, however, formed oligomers at 37°C. Fibrin-fibrinogen interactions were studied by affinity chromatography. At 37°C, des-A fibrin did not interact with immobilized fibrinogen, whereas des-AB fibrin stuck to it. Small amounts of des-A fibrin iv injected were cleared from the circulation much faster than des-AB fibrin. Large amounts of des-A fibrin injected into animals aggregated to microclots. These results suggest that des-A fibrin in a low concentration (0.01 mg/ml) does not form circulating oligomers and can very fast be cleared from the circulation possibly because of its weak interaction with fibrinogen. Aggregated des-A fibrin, however, is cleaved to form des-AB fibrin. Des-AB fibrin binds fibrinogen making it accessible to further thrombin cleavage. Thus, the threshold of clot formation would be the concentration of des-A fibrin in plasma. The preferential sequence of reactions is: fibrinogen → des-A fibrin → des-A fibrin aggregates → des-AB fibrin aggregates. Des-AB fibrin + fibrinogen → des-AB fibrin-fibrinogen → des-AB fibrin-des-A fibrin etc.

Applicability of Instability Index for In vitro Protein Stability Prediction

2019 ◽  
Vol 26 (5) ◽  
pp. 339-347 ◽  
Author(s):  
Dilani G. Gamage ◽  
Ajith Gunaratne ◽  
Gopal R. Periyannan ◽  
Timothy G. Russell
Keyword(s):  
Protein Stability ◽  
Caulobacter Crescentus ◽  
Effect Of Temperature ◽  
Instability Index ◽  
Dipeptide Composition ◽  
Experimental Conditions ◽  
The Stability ◽  
Vitro Protein

Background: The dipeptide composition-based Instability Index (II) is one of the protein primary structure-dependent methods available for in vivo protein stability predictions. As per this method, proteins with II value below 40 are stable proteins. Intracellular protein stability principles guided the original development of the II method. However, the use of the II method for in vitro protein stability predictions raises questions about the validity of applying the II method under experimental conditions that are different from the in vivo setting. Objective: The aim of this study is to experimentally test the validity of the use of II as an in vitro protein stability predictor. Methods: A representative protein CCM (CCM - Caulobacter crescentus metalloprotein) that rapidly degrades under in vitro conditions was used to probe the dipeptide sequence-dependent degradation properties of CCM by generating CCM mutants to represent stable and unstable II values. A comparative degradation analysis was carried out under in vitro conditions using wildtype CCM, CCM mutants and two other candidate proteins: metallo-β-lactamase L1 and α -S1- casein representing stable, borderline stable/unstable, and unstable proteins as per the II predictions. The effect of temperature and a protein stabilizing agent on CCM degradation was also tested. Results: Data support the dipeptide composition-dependent protein stability/instability in wt-CCM and mutants as predicted by the II method under in vitro conditions. However, the II failed to accurately represent the stability of other tested proteins. Data indicate the influence of protein environmental factors on the autoproteolysis of proteins. Conclusion: Broader application of the II method for the prediction of protein stability under in vitro conditions is questionable as the stability of the protein may be dependent not only on the intrinsic nature of the protein but also on the conditions of the protein milieu.

The effect of temperature and time on the properties of 2D Cs2ZnBr4 perovskite nanocrystals and its application in a Schottky barrier device

2021 ◽  
Author(s):  
Olusola Akinbami ◽  
Grace N Ngubeni ◽  
Francis Otieno ◽  
Rudo Kadzutu-Sithole ◽  
Cebisa Linganiso ◽  
...  
Keyword(s):  
Solar Cell ◽  
Schottky Barrier ◽  
Effect Of Temperature ◽  
Inorganic Perovskite ◽  
Perovskite Nanocrystals ◽  
The Stability ◽  
Barrier Device

2D hybrid perovskites are promising materials for solar cell applications, in particular, cesium based perovskite nanocrystals as they offer the stability that is absent in organic-inorganic perovskite. However, the most...

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