An Evaluation of a Plasma Fractionation System

1960 ◽  
Vol 4 (01) ◽  
pp. 031-044
Author(s):  
George Y. Shinowara ◽  
E. Mary Ruth
Keyword(s):  
Biological Activity ◽  
Ionic Strength ◽  
Plasma Proteins ◽  
High Concentration ◽  
Significant Evidence ◽  
Native Proteins ◽  
Mobility Data ◽  
Fractionation Procedure ◽  
The Individual ◽  
Low Ionic Strength

SummaryFour primary fractions comprising at least 97 per cent of the plasma proteins have been critically appraised for evidence of denaturation arising from a low temperature—low ionic strength fractionation system. The results in addition to those referable to the recovery of mass and biological activity include the following: The high solubilities of these fractions at pH 7.3 and low ionic strengths; the compatibility of the electrophoretic and ultracentrifugal data of the individual fractions with those of the original plasma; and the recovery of hemoglobin, not hematin, in fraction III obtained from specimens contaminated with this pigment. However, the most significant evidence for minimum alterations of native proteins was that the S20, w and the electrophoretic mobility data on the physically recombined fractions were identical to those found on whole plasma.The fractionation procedure examined here quantitatively isolates fibrinogen, prothrombin and antithrombin in primary fractions. Results have been obtained demonstrating its significance in other biological systems. These include the following: The finding of 5 S20, w classes in the 4 primary fractions; the occurrence of more than 90 per cent of the plasma gamma globulins in fraction III; the 98 per cent pure albumin in fraction IV; and, finally, the high concentration of beta lipoproteins in fraction II.

Aminoacyl-tRNA synthetases of Halobacterium cutirubrum: preliminary fractionation studies

1970 ◽  
Vol 48 (8) ◽  
pp. 944-946 ◽  
Author(s):  
E. Griffiths
Keyword(s):  
Ionic Strength ◽  
Gel Filtration ◽  
Halophilic Bacterium ◽  
Trna Synthetase ◽  
Partial Purification ◽  
Trna Synthetases ◽  
Aminoacyl Trna Synthetases ◽  
Fractionation Procedure ◽  
The Stability ◽  
Low Ionic Strength

The stability, in solutions of low ionic strength, of aminoacyl-tRNA synthetases from the extremely halophilic bacterium Halobacterium cutirubrum was studied as a preliminary to their fractionation. The enzymes differed considerably in their sensitivity to such solutions. Conditions were found where reactivation from the salt-free and inactive state could be achieved. Removal of both K+ and Mg2+ together generally resulted in better stability than the removal of K+ alone. A low temperature (4°) was also important for stability in buffers of low ionic strength. In some cases the L-amino acid substrates afforded protection against inactivation in the salt-free state. Gel filtration in low ionic strength medium was found to work well as a fractionation procedure; a partial purification of phenylalanyl-tRNA synthetase was effected in this way. The use of other conventional protein fractionation procedures is now possible.

scholarly journals Some electron-transfer reactions involving carbodi-imide-modified cytochrome c

1987 ◽  
Vol 243 (2) ◽  
pp. 379-384 ◽  
Author(s):  
A J Mathews ◽  
T Brittain
Keyword(s):  
Electron Transfer ◽  
Ionic Strength ◽  
Protein Charge ◽  
Native Proteins ◽  
Wide Range ◽  
Internal Electron ◽  
Modified Proteins ◽  
Time Courses ◽  
Low Ionic Strength ◽  
Kinetics Of

The reaction kinetics of native and carbodi-imide-modified tuna and horse heart cytochromes c with both a strong (dithionite) and a relatively weak (ascorbate) reducing agent were studied over a wide range of conditions. In their reactions with dithionite both the native and modified cytochromes exhibit single exponential time courses. The effects of dithionite concentration and ionic strength on the rate of the reduction are complex and can best be explained in terms of the model proposed by Lambeth & Palmer [(1973) J. Biol. Chem. 248, 6095-6103]. According to this model, at low ionic strength the native proteins are reduced almost exclusively by S2O4(2-) whereas the modified proteins showed reactivity towards both S2O4(2-) and SO2.-. These findings are interpreted in terms of the different charge characteristics of the carbodi-imide-modified proteins relative to the native proteins. The findings that the modified proteins react with ascorbate in a biphasic manner are explained as arising from ascorbate binding to a reducible form of the protein, before electron transfer, with an equilibrium between the ascorbate-reducible form of the protein and a non-reducible form. Estimates were obtained for both the ascorbate equilibrium binding constant and the rate constant for the internal electron transfer for both the native and modified horse and tuna proteins. The effect of pH on the reactions indicates that the active reductant in all cases is ascorbate2-. The studies of ascorbate reactivity yield important information concerning the proposed correlation between ascorbate reducibility and the presence of a 695 nm-absorption band, and the study of dithionite reactivity illustrates the effect of protein charge and solution ionic strength on the relative contributions made by the species SO2.- and S2O4(2-) to the reduction of ferricytochrome c.

CHROMATOGRAPHIC SEPARATION OF PHOSVITIN, α- AND β-LIPOVITELLIN OF EGG YOLK GRANULES ON TEAE-CELLULOSE

1964 ◽  
Vol 42 (8) ◽  
pp. 1203-1215 ◽  
Author(s):  
M. W. Radomski ◽  
W. H. Cook
Keyword(s):  
Ionic Strength ◽  
Egg Yolk ◽  
Gradient Elution ◽  
Partial Recovery ◽  
Linear Gradient ◽  
Yolk Granules ◽  
Sedimentation Behavior ◽  
Strength Limit ◽  
The Individual ◽  
Low Ionic Strength

The two components of lipovitellin and the three major components of yolk granules, phosvitin, α- and β-lipovitellin, have been separated by gradient elution chromatography on TEAE-cellulose. A 0.2 M phosphate buffer (pH 6.8) had the necessary ionic strength to dissolve these proteins and when applied in this solvent all components except β-lipovitellin were retained by the column. A linear gradient of ionic strength (limit buffer 0.2 M phosphate plus 0.5 M NaCl) was used to remove the other components. Recovery was essentially complete and the composition and properties of the individual components were similar to those obtained by previous chromatographic methods that gave only partial recovery. An additional component eluted after α-lipovitellin and before phosvitin, previously observed in Dowex-1 separations, was also observed by the present method. The composition, sedimentation behavior, and absorption spectra of this component indicate that it is a soluble complex of phosvitin and lipovitellin. When granules are dissolved in alkaline solvents (pH 9.4) of low ionic strength (0.05), phosvitin is not evident as a separate component during ultracentrifugation, but appears as the ionic strength is increased.

Isolation of Antithrombin Globulin from Human Blood Plasma

1960 ◽  
Vol 4 (01) ◽  
pp. 017-030
Author(s):  
George Y. Shinowara ◽  
Dimite J. Buckley
Keyword(s):  
Ionic Strength ◽  
Test Procedure ◽  
Human Blood Plasma ◽  
First Order ◽  
Antithrombin Activity ◽  
Low Concentrations ◽  
Fractionation Procedure ◽  
The Mean ◽  
Low Ionic Strength ◽  
Kinetics Of

SummaryFraction III + IV obtained by a low temperature — low ionic strength fractionation procedure was found to contain 97 percent of the antithrombin activity of whole plasma. The mean antithrombin activity in this fraction from 88 normal plasma specimens was 172.6 ± S. D. 19.8 units per ml. Further purification experiments resulted in fraction III representing 96 per cent of the antithrombin activity in fraction III + IV. There was no evidence for fibrinogen, prothrombin and antihemophilic globulin contamination in fraction III. Over 90 per cent of the plasma gamma globulins was present in this fraction.The kinetics of the thrombin-antithrombin reaction were investigated. Under the specific conditions employed, the reaction was found to follow a first order course. The heat of activation was determined to be 11,300 calories. A standard antithrombin unit is defined. A quantitative test procedure which is accurate at both high and low concentrations of the circulating anticoagulant is described and its application on fractions, heat defibrinated plasma and serum is discussed.

scholarly journals Chromatography of plasma proteins on immobilized Cibacron Blue F3-GA. Mechanism of the molecular interaction

1982 ◽  
Vol 203 (3) ◽  
pp. 637-641 ◽  
Author(s):  
E Gianazza ◽  
P Arnaud
Keyword(s):  
Molecular Weight ◽  
Ionic Strength ◽  
Molecular Interaction ◽  
Plasma Proteins ◽  
Hydrophobic Interactions ◽  
Molecular Characteristics ◽  
Acidic Ph ◽  
Cibacron Blue ◽  
Ligand Affinity ◽  
Low Ionic Strength

Fractionation of plasma proteins on immobilized Cibacron Blue F3-GA (Affi-gel Blue) under different conditions of pH, ionic strength and temperature was studied. At acidic pH the unbound proteins were eluted in order of increasing pI (the Affi-gel Blue behaving as ion-exchanger); at basic pH and at low ionic strength they were eluted in order of decreasing molecular weight (separation by diffusion-exclusion). For the proteins that were either retarded in comparison with substances of similar molecular characteristics, or that were bound to the resin, pseudo-ligand affinity or hydrophobic interactions were also implicated.

scholarly journals The participation of phospholipids in the interaction of leucocidin and the cell membrane of the polymorphonuclear leucocyte

1967 ◽  
Vol 105 (3) ◽  
pp. 1029-1038 ◽  
Author(s):  
A. M. Woodin ◽  
Antonnette A. Wieneke
Keyword(s):  
Biological Activity ◽  
Fatty Acid ◽  
Light Scattering ◽  
Ionic Strength ◽  
Cell Membrane ◽  
Phosphatidic Acid ◽  
Polymorphonuclear Leucocyte ◽  
Side Chains ◽  
Low Ionic Strength

1. The interaction of the two components of leucocidin with various lipids has been studied by sedimentation, flotation, light-scattering and changes in the biological activity of leucocidin. 2. Phosphatidylserine, phosphatidylcholine, diphosphoinositide, triphosphoinositide and phosphatidic acid, but not phosphatidylethanolamine, lysophosphatidylcholine, cerebrosides, gangliosides or tristearin, induce aggregation of the F component of leucocidin. 3. The S component of leucocidin does not interact directly with these phospholipids, but interacts with the F component of leucocidin after its modification by lipids. 4. The increased sedimentation or light-scattering induced by low phospholipid concentrations is reversed at higher phospholipid concentrations. 6. The aggregates formed by phospholipids and leucocidin are due, not to adsorption of leucocidin alone, but also to the formation of leucocidin polymers. 7. It is concluded that the aggregation is due to the interaction of the F component with the fatty acid side chains in the lipid micelle. 8. The S component of leucocidin is inactivated by triphosphoinositide at physiological ionic strength; the F component of leucocidin is inactivated at low ionic strength by triphosphoinositide and remains inactive when the ionic strength is increased. 9. It is suggested that in the leucocyte cell membrane the S component of leucocidin interacts with the polar hydrophilic groups of triphosphoinositide and that the F component of leucocidin interacts with the hydrophobic parts of triphosphoinositide.

scholarly journals Interaction Between Rabbit Erythroblast Ferritin and Normal Plasma Proteins

Blood ◽  
1974 ◽  
Vol 43 (6) ◽  
pp. 875-883 ◽  
Author(s):  
Hideo Yamada ◽  
Thomas G. Gabuzda ◽  
Marion Berenfeld
Keyword(s):  
Ionic Strength ◽  
Guinea Pig ◽  
Plasma Proteins ◽  
Starch Granule ◽  
Binding Activity ◽  
The Other ◽  
Heat Stable ◽  
Normal Plasma ◽  
Low Ionic Strength ◽  
Plasma Factors

Abstract Erythroblast ferritin (EF), isolated from phenylhydrazine-anemic rabbit marrow, interacts with components of normal plasma and forms a complex separable by starch granule electrophoresis and by dialysis techniques. The binding is facilitated at low ionic strength. The plasma factors responsible for binding EF are present in autologous, as well as homologous, plasma of normal nonimmune rabbits, although binding activities are quite variable in different plasmas. The binding activity for rabbit EF is also present in heterologous plasma of mouse, guinea pig, and man. The active principles in plasma were identified as two heat-stable components which fractionate as immunoglobulins, one as IgG and the other as IgM. The results support the hypothesis that natural antiferritin antibodies of low titer are present in normal plasma.

Conformational Transitions in Escherichia coli Ribosomal RNAs as Visualized by Dedicated STEM

1988 ◽  
Vol 46 ◽  
pp. 176-177
Author(s):  
J.S. Wall ◽  
V. Maridiyan ◽  
S. Tumminia ◽  
J. Hairifeld ◽  
M. Boublik
Keyword(s):  
Ionic Strength ◽  
Three Dimensional ◽  
Conformational Transitions ◽  
Dark Field ◽  
Dimensional Structure ◽  
Ribosomal Rnas ◽  
Field Mode ◽  
Freeze Dried ◽  
High Resolution Images ◽  
Low Ionic Strength

The high contrast in the dark-field mode of dedicated STEM, specimen deposition by the wet film technique and low radiation dose (1 e/Å2) at -160°C make it possible to obtain high resolution images of unstained freeze-dried macromolecules with minimal structural distortion. Since the image intensity is directly related to the local projected mass of the specimen it became feasible to determine the molecular mass and mass distribution within individual macromolecules and from these data to calculate the linear density (M/L) and the radii of gyration.2 This parameter (RQ), reflecting the three-dimensional structure of the macromolecular particles in solution, has been applied to monitor the conformational transitions in E. coli 16S and 23S ribosomal RNAs in solutions of various ionic strength.In spite of the differences in mass (550 kD and 1050 kD, respectively), both 16S and 23S RNA appear equally sensitive to changes in buffer conditions. In deionized water or conditions of extremely low ionic strength both appear as filamentous structures (Fig. la and 2a, respectively) possessing a major backbone with protruding branches which are more frequent and more complex in 23S RNA (Fig. 2a).

Calculation of Predictive Odds for Possible Carriers of Haemophilia

1975 ◽  
Vol 34 (03) ◽  
pp. 740-747 ◽  
Author(s):  
C. R. M Prentice ◽  
C. D Forbes ◽  
Sandra Morrice ◽  
A. D McLaren
Keyword(s):  
Biological Activity ◽  
Factor Viii ◽  
Factor Viii Related Antigen ◽  
Using Data ◽  
The Individual ◽  
Betting Odds

SummaryBetting odds for possible carriers of haemophilia have been calculated using data derived from normal and known carrier populations. For each possible carrier the concentration of factor VIII-related antigen and factor VIII biological activity was measured and used to determine the probability of the individual being a carrier. The calculations indicated that, of the 32 possible carriers, 11 were likely to be normal (odds of more than 5:1) while 11 were likely to be haemophilia carriers (again odds of more than 5:1).

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