scholarly journals Subcellular structure of bovine thyroid gland. A study on bovine thyroid membranes by buoyant-density-gradient centrifugation in a B-XIV zonal rotor

1975 ◽  
Vol 152 (3) ◽  
pp. 601-607 ◽  
Author(s):  
H J Hilderson ◽  
M J De Wolf ◽  
A R Lagrou ◽  
W S Dierick
Keyword(s):  
Endoplasmic Reticulum ◽  
Thyroid Gland ◽  
Density Gradient ◽  
Rough Endoplasmic Reticulum ◽  
Plasma Membranes ◽  
Smooth Endoplasmic Reticulum ◽  
Gradient Centrifugation ◽  
Buoyant Density ◽  
Bovine Thyroid ◽  
Zonal Rotor

A combined mitochondrial and light mitochondrial fraction and a microsomal fraction were isolated from bovine thyroid gland and fractionated further in a B-XIV zonal rotor. A density gradient ranging from 20 to 50% (w/w) sucrose was used. The rotor was operated for 3 h at 45 000 rev./min. All manipulations were performed at 4 degrees C and at pH 7.4. 2. Membranous material was recovered in two zones: zone I, containing microsomal material derived from both smooth endoplasmic reticulum and plasma membranes and probably also from other smooth membranes; zone II, containing material from rough endoplasmic reticulum. 3. Increasing the pH of the medium up to 8.6, or the addition of Mg2+ to the medium resulted in the formation of a single zone at intermediate densities (aggregation of membranes?). An analogous effect was obtained after treatment with Pb (NO3) 2. 4. In the presence of heparin (50 i.u./ml) the bulk of the membranes was found in zone I. This was due to the release of ribosomes from the rough endoplasmic reticulum.

Les systèmes membranaires de Scopulariopsis brevicaulis. II. Étude comparée de leur composition lipidique

1987 ◽  
Vol 33 (1) ◽  
pp. 78-81 ◽  
Author(s):  
Jean-Louis Fonvieille ◽  
Jane-Marie Touze-Soulet ◽  
Charles Montant
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membranes ◽  
Smooth Endoplasmic Reticulum ◽  
Gradient Centrifugation ◽  
Percoll Gradient ◽  
Mitochondrial Membranes ◽  
Isolation Method ◽  
Discontinuous Sucrose Gradient ◽  
Phospholipid Ratio ◽  
Scopulariopsis Brevicaulis

Biochemical studies of Scopulariopsis brevicaulis membranes (except mitochondrial membranes), in particular their lipid composition, were undertaken using plasma membranes and fractions containing smooth endoplasmic reticulum and Golgi apparatus membranes. The results obtained by two separation techniques, a continuous Percoll gradient and a discontinuous sucrose gradient centrifugation, were similar. Depending on the isolation method used, the plasma membrane contained 30 or 33% protein and 23 or 30% lipid of which 16.5 or 18% was phospholipid. The sterol/phospholipid ratio was 1.98 or 2.28. The endoplasmic reticulum + Golgi fraction contained 21 or 26.7% protein and 43 or 50% lipid of which 7 or 10% was phospholipid. The sterol/phospholipid ratio was 0.85 or 0.81.

scholarly journals Electrophoretic mobility of γ-glutamyltransferase in rat liver subcellular fractions. Evidence for structure difference from the kidney enzyme

1989 ◽  
Vol 262 (2) ◽  
pp. 535-539 ◽  
Author(s):  
B Antoine ◽  
A Visvikis ◽  
C Thioudellet ◽  
A Rahimi-Pour ◽  
N Strazielle ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Liver Enzyme ◽  
Rat Liver ◽  
Rough Endoplasmic Reticulum ◽  
Plasma Membranes ◽  
Smooth Endoplasmic Reticulum ◽  
Rat Kidney ◽  
Immunoblot Analysis ◽  
Subcellular Fractions ◽  
Golgi Membranes

Adult rat liver gamma-glutamyltransferase (GGT) has been poorly characterized because of its very low concentration in the tissue. In contrast with the kidney, the liver enzyme is inducible by some xenobiotics, and its relationship to hepatic ontogeny and carcinogenesis seems to be important. Liver GGT polypeptides were identified by immunoblot analysis in subcellular fractions (rough endoplasmic reticulum, smooth endoplasmic reticulum, Golgi membranes and plasma membranes). Rat liver GGT appeared as a series of polypeptides corresponding to different maturation steps. Polypeptides related to the heavy subunit of GGT were detected in rough endoplasmic reticulum at 49, 53 and 55 kDa, and in Golgi membranes at 55, 60 and 66 kDa. Two polypeptides related to the light subunit of GGT were also observed in Golgi membranes. In plasma membranes GGT was composed of 100 kDa, 66 kDa and 31 kDa polypeptides. The 66 kDa component could correspond to the heavy subunit of the rat liver enzyme, and if so has a molecular mass higher than that of the purified rat kidney form of GGT (papain-treated). These data suggest different peptide backbones for the heavy subunits of liver GGT and kidney GGT.

The presence of epidermal growth factor binding sites in the intracellular organelles of term human placenta

1986 ◽  
Vol 84 (1) ◽  
pp. 19-40
Author(s):  
N. Ramani ◽  
N. Chegini ◽  
C.V. Rao ◽  
P.G. Woost ◽  
G.S. Schultz
Keyword(s):  
Endoplasmic Reticulum ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Rough Endoplasmic Reticulum ◽  
Human Placenta ◽  
Binding Sites ◽  
Plasma Membranes ◽  
Smooth Endoplasmic Reticulum ◽  
Intracellular Organelles ◽  
Epidermal Growth

Highly purified lysosomes, rough and smooth endoplasmic reticulum, and Golgi apparatus, as well as microvillus plasma membranes, bound 125I-labelled epidermal growth factor ([125I]EGF) with similar affinity. Scatchard plots for all the organelles were curvilinear. The apparent number of available binding sites per mg protein of intracellular organelles was 27–71% of that found in microvillus plasma membranes. The bound and free [125I]EGF were not degraded by any of the organelles. Binding and dissociation of [125I]EGF in all organelles were dependent on the time and temperature of incubation. The specificity of [125I]EGF binding was similar in all organelles. The optimal pH for binding to lysosomes was 6.0, in contrast to 7.0 for all the other organelles. Exposure of different organelles to enzymes and protein-modifying reagents resulted in numerous binding differences between the intracellular organelles and microvillus plasma membranes. Covalent affinity labelling with [125I]EGF revealed two major proteins of 155 and 140(X10(3)) Mr in all the organelles. The 155 X 10(3) Mr protein was labelled predominantly in all organelles except rough endoplasmic reticulum, where both proteins were equally labelled. Addition of proteolytic inhibitors during isolation of organelles did not alter the pattern of [125I]EGF-labelled binding proteins found in the organelles. EGF also stimulated phosphorylation of the 155 and 140(X10(3)) Mr proteins in all the organelles. The 155 X 10(3) Mr protein was phosphorylated more than the 140 X 10(3) Mr protein in microvillus plasma membranes and smooth endoplasmic reticulum, whereas the 140 X 10(3) Mr protein was phosphorylated more than the 155 X 10(3) Mr protein in lysosomes and both proteins were equally phosphorylated in rough endoplasmic reticulum. Several organelles also contained minor [125I]EGF-binding proteins that did not show phosphorylation response and proteins that showed phosphorylation response but did not bind [125I]EGF. Thus, the present study demonstrates by a number of different criteria, that several intracellular organelles of term human placenta also contain EGF-binding and kinase activities.

scholarly journals Glycosyltransferase activities in Golgi complex and endoplasmic reticulum fractions isolated from African trypanosomes.

1984 ◽  
Vol 99 (2) ◽  
pp. 569-577 ◽  
Author(s):  
D J Grab ◽  
S Ito ◽  
U A Kara ◽  
L Rovis
Keyword(s):  
Endoplasmic Reticulum ◽  
Rough Endoplasmic Reticulum ◽  
Golgi Complex ◽  
Smooth Endoplasmic Reticulum ◽  
Surface Glycoprotein ◽  
Relative Distribution ◽  
African Trypanosomes ◽  
Independent Form ◽  
Variable Surface ◽  
Dependent Form

Highly enriched Golgi complex and endoplasmic reticulum fractions were isolated from total microsomes obtained from Trypanosoma brucei, Trypanosoma congolense, and Trypanosoma vivax, and tested for glycosyltransferase activity. Purity of the fractions was assessed by electron microscopy as well as by biochemical analysis. The relative distribution of all the glycosyltransferases was remarkably similar for the three species of African trypanosomes studied. The Golgi complex fraction contained most of the galactosyltransferase activity followed by the smooth and rough endoplasmic reticulum fractions. The dolichol-dependent mannosyltransferase activities were highest for the rough endoplasmic reticulum, lower for the smooth endoplasmic reticulum, and lowest for the Golgi complex. Although the dolichol-independent form of N-acetylglucosaminyltransferase was essentially similar in all the fractions, the dolichol-dependent form of this enzyme was much higher in the endoplasmic reticulum fractions than in the Golgi complex fraction. Inhibition of this latter activity in the smooth endoplasmic reticulum fraction by tunicamycin A1 suggests that core glycosylation of the variable surface glycoprotein may occur in this organelle and not in the rough endoplasmic reticulum as previously assumed.

The isolation of plasma membrane from protoplasts of soybean suspension cultures

1977 ◽  
Vol 24 (1) ◽  
pp. 295-310
Author(s):  
D.W. Galbraith ◽  
D.H. Northcote
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Membrane Fraction ◽  
Sucrose Gradient ◽  
Plasma Membranes ◽  
Gradient Centrifugation ◽  
Buoyant Density ◽  
Membrane Systems ◽  
Enzymic Digestion ◽  
Nadh Cytochrome

A procedure for the isolation of plasma membranes from protoplasts of suspension-cultured soybean is described. Protoplasts were prepared by enzymic digestion of the cell wall and the plasma membrane was labelled with radioactive diazotized sulphanilic acid. The membrane systems from broken protoplasts were separated by continuous isopycnic sucrose gradient centrifugation. Radioactivity was localized in a band possessing a buoyant density of 1–14 g ml-1. The activities of NADPH- and NADH-cytochrome c reductase, fumarase, Mg2+-ATPase, IDPase and acid phosphodiesterase in the various regions of the density gradient were determined. A plasma membrane fraction was selected which was relatively uncontaminated with membranes derived from endoplasmic reticulum, tonoplasts and mitochondria. The results indicated that Mg2+-ATPase and possibly acid phosphodiesterase were associated with the plasma membrane.

scholarly journals Evidence that platelet density depends on the alpha-granule content in platelets

Blood ◽  
1984 ◽  
Vol 63 (2) ◽  
pp. 482-485 ◽  
Author(s):  
BA van Oost ◽  
AP Timmermans ◽  
JJ Sixma
Keyword(s):  
Density Distribution ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Platelet Rich Plasma ◽  
Gradient Centrifugation ◽  
Buoyant Density ◽  
Adenosine Diphosphate ◽  
Density Gradients

Abstract The relation between platelet buoyant density and beta-thromboglobulin (beta-TG), a marker for platelet alpha-granule content, was assessed by three independent approaches. (1) Platelets were separated on iso- osmolar discontinuous Stractan density gradients into five fractions, ranging in density from 1.061 g/ml to 1.091 g/ml (20 degrees C). The beta-TG content (mean +/- SD, n = 17) increased with the platelet density from 27.8 +/- 8.6 micrograms beta-TG/10(9) cells (20% less- dense platelets) up to 65.6 +/- 15.5 micrograms beta-TG/10(9) cells (15% most-dense platelets). (2) Activation of platelets in platelet- rich plasma with thrombin, adenosine diphosphate, collagen, or epinephrine resulted in a decreased density of the platelets. This was only seen when there was simultaneous secretion of beta-TG. (3) The less-dense and the more-dense platelet fractions, after isolation by density gradient centrifugation, were separately treated with thrombin. After complete degranulation, the density distribution of the originally less-dense and more-dense platelets were identical and were much narrower than the density distribution of resting platelets.

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