Rapid release of peroxidase by peanut cells in suspension culture

1973 ◽  
Vol 51 (6) ◽  
pp. 1169-1175 ◽  
Author(s):  
R. B. van Huystee ◽  
G. Turcon
Keyword(s):  
Molecular Weight ◽  
Metabolic Rate ◽  
Suspension Culture ◽  
Gel Filtration ◽  
Electron Microscopic ◽  
Rapid Release ◽  
Microscopic Studies ◽  
Very High

Peanut cells in a suspension culture were studied with regard to the process of peroxidase release and some ultrastructural features that appeared to relate to this process. By means of gel filtration on Sephadex G-200 it was established that protein released into the medium had the same molecular weight as the peroxidase found in this medium. From information gained through the electron-microscopic studies and from incorporation of labeled leucine, it may be deduced that the metabolic rate of these cells is very high. Within 2 h after incubation with radioactive leucine it was noted that newly synthesized protein was released by the cells into the medium. This protein had also the same characteristics of peroxidase and its synthesis and subsequent appearance in the medium could be reduced by incubation in the presence of cycloheximide. A possible explanation for the presence of these large quantities of peroxidase is discussed.

scholarly journals Biosynthesis of glucagon in isolated pancreatic islets of guinea pigs

1974 ◽  
Vol 140 (1) ◽  
pp. 13-21 ◽  
Author(s):  
Claes Hellerström ◽  
Simon L. Howell ◽  
John C. Edwards ◽  
Arne Andersson ◽  
Claes-Göran Östenson
Keyword(s):  
Molecular Weight ◽  
Biological Activity ◽  
Gel Filtration ◽  
Subcellular Fractionation ◽  
Fat Cell ◽  
Electron Microscopic ◽  
Isolated Pancreatic Islets ◽  
Isolated Islets Of Langerhans ◽  
Electron Microscopic Radioautography ◽  
Very High

1. The biosynthesis of glucagon in guinea-pig A2 cells was investigated by incubation of isolated islets of Langerhans in the presence of [3H]tryptophan for periods of up to 14 days. Proteins were extracted from islets and incubation media and analysed by gel filtration. 2. In addition to very-high-molecular-weight (100000) proteins, the principal tryptophan-containing biosynthetic product after incubation for up to 17h was a protein of minimum mol.wt. 9000, which co-eluted on gel filtration with a peak of glucagon-like immunoreactivity, but was apparently devoid of biological activity in a fat-cell assay. A discrete peak of labelled glucagon was only recovered after incubation for at least 6 days. Losses of glucagon during the extraction and rapid secretion of newly synthesized glucagon into incubation media were excluded as reasons for the lack of recovery of labelled hormone from islets after shorter incubations. 3. The 9000-mol.wt. protein was localized to A2 cells in experiments using B-cell-depleted islets, and to A2-cell granules by subcellular fractionation and electron-microscopic radioautography. Only glucagon was secreted into the incubation medium. 4. Possible relationships between the 9000-mol.wt. protein and glucagon are discussed in the light of postulated mechanisms of glucagon biosynthesis.

Estramustine binds MAP-2 to inhibit microtubule assembly in vitro

1988 ◽  
Vol 89 (3) ◽  
pp. 331-342
Author(s):  
M.E. Stearns ◽  
K.D. Tew
Keyword(s):  
Linear Regression Analysis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Microtubule Assembly ◽  
Scatchard Analysis ◽  
Binding Assays ◽  
Electron Microscopic ◽  
Associated Proteins ◽  
Microscopic Studies

We have investigated the ability of estramustine to bind to rat brain microtubule-associated proteins (MAPs) and purified MAP-2 in vitro. [3H]estramustine's relative affinity for tubulin and MAPs was assessed by gel filtration chromatography, immunoprecipitation and binding assays. Scatchard analysis demonstrated a specific affinity of the drug for MAP-2. Calculations from kinetic parameters and non-linear regression analysis gave a Kd of 15 microM, and a Bmax of 3.4 × 10(−7)M ml-1. Extrapolation of this value suggested that each MAP-2 molecule binds approximately 20 molecules of estramustine. Microtubule assembly studies and SDS-polyacrylamide gel electrophoresis revealed that at 20–60 microM levels, estramustine inhibited the association of MAPs with taxol microtubules. Turbidity (A350) studies further demonstrated that 20–60 microM-estramustine inhibited MAP-2-driven tubulin assembly and produced microtubule disassembly. Electron-microscopic studies confirmed the centrifugation and turbidity results. The data demonstrated that estramustine can bind MAPs and MAP-2 specifically, thereby inhibiting microtubule assembly.

Distribution of radioactive silver in the subcellular fractions of various tissues of the rat and its binding to low molecular weight proteins

1983 ◽  
Vol 61 (11) ◽  
pp. 1391-1395 ◽  
Author(s):  
Yousef Matuk
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Gel Filtration ◽  
Total Radioactivity ◽  
Subcellular Fractions ◽  
Low Molecular Weight ◽  
Electron Microscopic ◽  
Molecular Weight Peak

In view of the electron microscopic evidence that silver does not penetrate cellular barriers, the distribution of radioactive silver in rat blood and subcellular fractions of liver, kidneys, spleen, and forebrain was studied. It was found that 24 h after a single intraperitoneal injection high levels of radioactivity were reached which decreased at different rates in the various tissues studied. In plasma, liver, and kidneys there was an initial rapid loss of radioactivity which was followed by a slower rate of loss. In the blood, forebrain, and spleen the loss of radioactivity was linear and somewhat slower than in the other three tissues. The cytosols of the liver and kidneys contained 60% while those of the forebrain and spleen contained 30% of the total radioactivity found in the tissue homogenates. Gel filtration on Sephadex G-75 showed that all cytosols contained two peaks of radioactivity; a high molecular weight peak which eluted just after the void volume and a low molecular weight peak. The amount of radioactivity in both peaks was, however, much lower in the chromatographic peaks of the forebrain and spleen than that found in those of the liver and kidneys. Furthermore, the spleen had a comparatively very small low molecular weight radioactive peak. In vitro experiments with liver cytosol showed similar results to those found in vivo in that the high molecular weight radioactive peak could be removed by heat. It is concluded that silver does enter cells and that silver thionein exists in the cytosols of forebrain, spleen, kidney, and liver.

scholarly journals Size heterogeneity of rat pituitary prolactin

1980 ◽  
Vol 189 (3) ◽  
pp. 605-614 ◽  
Author(s):  
M Wallis ◽  
M Daniels ◽  
S A Ellis
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Paper Electrophoresis ◽  
Freezing And Thawing ◽  
Molecular Weights ◽  
Size Heterogeneity ◽  
Very High

The occurrence of multiple forms of rat prolactin with different molecular weights (size heterogeneity) was studied with anterior pituitary extracts, purified rat prolactin and 125I-labelled rat prolactin. In each case, three main forms of the hormone were detected by gel filtration on Sephadex G-100: a major one (80–90%) corresponding to monomeric prolactin (mol.wt. 22000–25000), a peak (8–20%) that could be a dimer (mol.wt. 45000–50000) and a small quantity (1–5%) of a component of much greater molecular weight. On freezing and thawing of 125I-labelled rat prolactin, there was little interconversion of monomer and ‘dimer’ peaks, but both were converted substantially to very high-molecular-weight material. All three peaks of 125I-labelled rat prolactin could be precipitated by anti-(rat prolactin) serum and all three gave similar patterns of radioactive peptides after digestion with chymotrypsin followed by high-voltage paper electrophoresis. On sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, the monomer peak of 125I-labelled prolactin migrated as a single component of mol.wt. 22000, the very high-molecular-weight peak largely dissociated to a component running in the same position as the monomer, and the ‘dimer’ peak migrated partly as a component of mol.wt. 45000 and partly as a component migrating with monomeric prolactin. No treatment was found that could dissociate the ‘dimer’ peak completely to monomeric prolactin.

scholarly journals The separation and distribution of simple and condensed leucoanthocyanins of the tea plant (Camellia sinensis L.)

1969 ◽  
Vol 113 (5) ◽  
pp. 757-763 ◽  
Author(s):  
G. I. Forrest ◽  
D S Bendall
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Inverse Correlation ◽  
Tea Plant ◽  
Degree Of Polymerization ◽  
Monomer Ratio ◽  
Lower Mobility ◽  
High Concentrations ◽  
Root Tissues ◽  
Very High

1. Leucoanthocyanin monomers of high mobilities in aqueous solvents on thinlayer chromatograms, assumed to be structurally simple, were characteristic of mature bulky tissues, whereas members of lower mobility were confined to young vegetative and floral tissues. 2. Flavylogens were separated by gel filtration on Sephadex columns into monomeric, oligomeric and polymeric fractions. 3. The polymeric fraction from young brown stems was heterogeneous, one-half having a molecular weight of about 3400, one-third a molecular weight between 3600 and 17000, and the remainder a molecular weight of over 17000. 4. Leaves had low flavylogen concentrations; only monomers were present. Stem tissues were rich in polymers, which increased with the age of the young stem and decreased inwards through the wood. The maximal flavylogen concentrations were in the phloem and cambium from mature stems, where all three fractions were richly present. The periderm tissue and, to a lesser extent, the seed coat were characterized by a very high polymer/monomer ratio, exhibiting a much higher degree of polymerization than the wood. Root tissues contained high concentrations of monomers. 5. In general, there was an inverse correlation between the extent of polymerization and the complexity of the monomers present. 6. The results are in favour of the thesis that the function of the flavanols is, after polymerization to condensed tannins, to impregnate dead structural tissues and thereby to protect them from infection and decay.

scholarly journals Electron-microscopic and electrophoretic studies of bovine femoral-head cartilage proteoglycan fractions

1986 ◽  
Vol 240 (1) ◽  
pp. 41-48 ◽  
Author(s):  
D J Thornton ◽  
I A Nieduszynski ◽  
K Oates ◽  
J K Sheehan
Keyword(s):  
Electron Microscopy ◽  
Femoral Head ◽  
Gel Electrophoresis ◽  
Gradient Centrifugation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Mixed Salt ◽  
Electron Microscopic ◽  
Femoral Head Cartilage ◽  
Microscopic Studies

Proteoglycans (A1D1) extracted from bovine femoral-head cartilage were examined by electron microscopy using benzyldimethylammonium chloride as a spreading agent. The preparation contained a mixture of particles, some with a ‘beaded’ structure and a contiguous filamentous ‘tail’ at one end and others which appeared as round ‘blobs’, some of which also had filamentous tails. Previous electron-microscopic studies of proteoglycan monomers have indicated that their length distributions were apparently unimodal, a finding that contrasted with agarose/polyacrylamide-gel-electrophoresis results, which generally indicated two bands. In the present study proteoglycans isolated from the slowly migrating electrophoretic band were shown to be predominantly the larger molecules of beaded appearance, whereas the rapidly migrating proteoglycans were predominantly molecules with the ‘blob-like’ appearance. Gel-filtration, isopycnic-density-gradient-centrifugation and rate-zonal-centrifugation techniques were evaluated as means of proteoglycan fractionation by electron microscopy and agarose-gel electrophoresis. Rate-zonal centrifugation in mixed-salt gradients of caesium chloride/4 M-guanidinium chloride yielded the most effective fractionation.

scholarly journals Electron Microscopic Studies of the Factor VIII-Von Willebrand Factor Protein

1977 ◽  
Author(s):  
V. J. Marder ◽  
L. Tranqui-Pouit ◽  
G. Hudry-Clergeon ◽  
V. Atichartakarn ◽  
M. Suscillon
Keyword(s):  
Factor Viii ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Molecular Size ◽  
Axial Ratio ◽  
Molecular Entity ◽  
Electron Microscopic ◽  
Human Factor Viii ◽  
Microscopic Studies ◽  
Von Willebrand

Preparations of human Factor VIII which were homogemeous by SDS-polyacrylamide gel electrophoresis were examined by electron microscopy after negative staining with phosphotungstic acid. Most striking were large oval and circular forms, usually present in loose groupings of four or more molecules. Approximately 80% were oval-shaped and the mean axial dimensions of 167 such molecules, considered to be prolate ellipsoids, were 725 Å and 346 Å (axial ratio about 2rl). The 34 circular forms had a mean diameter of 545 Å and were considered to represent a separate though related molecular entity, either of spherical or oblate ellipsoidal shape. A variety of smaller forms was also present in these preparations including round molecules of 110–330 Å in diameter, irregular ovoid shapes with filamentous transformation and filaments of variable length and thickness. Based on calculations of volume relative to that of fibrinogen, the molecular weight of the predominant prolate ellipsoid form is about 3 million daltons, a value which is consistent with reported estimates of molecular size obtained by ultracentrifuge or gel filtration studies.

scholarly journals Isolation and characterization of lipid-protein particles containing platelet factor 3 released from human platelets

1982 ◽  
Vol 203 (1) ◽  
pp. 303-311 ◽  
Author(s):  
H Sandberg ◽  
L O Andersson ◽  
S Höglund
Keyword(s):  
Gel Filtration ◽  
Phospholipid Composition ◽  
Chemical Characterization ◽  
Human Platelets ◽  
Electron Microscopic ◽  
Platelet Factor ◽  
Isolation And Characterization ◽  
Microscopic Studies ◽  
Time Test ◽  
Protein Particles

Lipid-protein particles with platelet factor 3 measured by the Stypven clotting-time test [Hardisty & Hutton (1966) Br. J. Haematol. 12, 764-776] have been isolated from platelet-release supernatant. Starting material was washed platelets, which were released by treatment with collagen. Purification of the particles from other components in the release material was accomplished by gel filtration on Sepharose CL-4B followed by affinity chromatography on poly-L-lysine-Sepharose CL-4B gel. Chemical characterization showed that the particles were composed of 40% protein, 42% phospholipids, 13% cholesterol and 5% triacylglycerols. The phospholipid composition was 38% phosphatidylcholine, 25% phosphatidylethanolamine, 9% phosphatidylserine, 2% phosphatidic acid and 26% sphingomyelin. No carbohydrate was detected. Electron-microscopic studies revealed the presence of membranous particles with diameters between 70 and 170 nm.

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