scholarly journals Freeze-fracture cytochemistry: partition of glycophorin in freeze-fractured human erythrocyte membranes.

1982 ◽  
Vol 93 (2) ◽  
pp. 463-469 ◽  
Author(s):  
P P da Silva ◽  
M R Torrisi
Keyword(s):  
Human Erythrocyte ◽  
Binding Sites ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Transmembrane Protein ◽  
Freeze Fracture ◽  
Membrane Skeleton ◽  
Erythrocyte Membranes ◽  
Con A ◽  
Human Erythrocyte Membranes

Thin-section and critical-point-dried fracture-labeled preparations are used to determine the distribution and partition of glycophorin-associated wheat germ agglutinin (WGA) binding sites over protoplasmic and exoplasmic faces of freeze-fractured human erythrocyte membranes. Most wheat germ agglutinin binding sites are found over exoplasmic faces. Label is sparse over the protoplasmic faces. These results contrast with previous observations of the partition of band 3 component where biochemical analysis and fracture-label of concanavalin A (Con A) binding sites show preferential partition of this transmembrane protein with the protoplasmic face. Presence of characteristic proportions of WGA and Con A binding sites over each fracture face is interpreted to indicate the operation of a stochastic process during freeze-fracture. This process appears modulated by the relative expression of each transmembrane protein at either surface as well as by their association to components of the erythrocyte membrane skeleton.

Freeze-fracture Electron Microscopic observations on the effects of sulphydryl group reagents on human erythrocyte membranes

1990 ◽  
Vol 48 (3) ◽  
pp. 524-525 ◽  
Author(s):  
Gheorghe Benga ◽  
Anthony Brain ◽  
Victor I. Pop ◽  
John Wrigglesworth
Keyword(s):  
Human Erythrocyte ◽  
Freeze Fracture ◽  
Band 3 ◽  
Erythrocyte Membranes ◽  
Electron Microscopic ◽  
Experimental Conditions ◽  
Water Permeation ◽  
Human Erythrocyte Membranes ◽  
Erythrocyte Membrane Proteins

The intra-membrane particles (IMPs) observed on the fracture face of frozen erythrocyte membranes are thought to correspond primarily to “band 3” tetramers or dimers. Some recent studies correlating the inhibition of water diffusion in erythrocytes by p-chloromercuribenzene sulfonate (PCMBS) with the binding of 203Hg to erythrocyte membrane proteins has enabled band 3 and the polypeptides in band 4.5 to be identified as the proteins associated with the channels for water permeation in human erythrocytes. A further characterization of the effects of the incubation of human erythrocyte membranes with PCMBS and N-ethylmaleimide (NEM) has been performed as previously described. Experimental conditions have been previously described.A comparison was made of the appearance of freeze-etched membranes of control erythrocytes and erythrocytes with the sulphydryl reagents. It was found that on many of the control and NEM-treated cells, small (50-100 nm) elevated patches could be seen (Fig. 1, 2 and 3). These are present on both fracture and etch faces and are devoid of any intramembrane particles. The patch elevations were never observed in the membranes of PCMBS-treated cells (Fig. 4).

Structure of cytochalasins and cytochalasin B binding sites in human erythrocyte membranes

Biochemistry ◽  
1980 ◽  
Vol 19 (4) ◽  
pp. 679-683 ◽  
Author(s):  
Amrit L. Rampal ◽  
Harold B. Pinkofsky ◽  
Chan Y. Jung
Keyword(s):  
Human Erythrocyte ◽  
Binding Sites ◽  
Cytochalasin B ◽  
Erythrocyte Membranes ◽  
Human Erythrocyte Membranes

scholarly journals Subfractionation of cardiac sarcolemma with wheat-germ agglutinin

1989 ◽  
Vol 264 (3) ◽  
pp. 885-892 ◽  
Author(s):  
J H M Charuk ◽  
S Howlett ◽  
M Michalak
Keyword(s):  
Plasma Membrane ◽  
Binding Sites ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Freeze Fracture ◽  
Protein Composition ◽  
Tubular Structures ◽  
Electron Microscopic ◽  
Cardiac Sarcolemma ◽  
Inside Out

The properties of highly purified bovine cardiac sarcolemma subfractionated with the lectin, wheat-germ agglutinin (WGA) were studied. Two different membrane subfractions were isolated, one which was agglutinated in the presence of 1.0 mg of WGA/mg of protein (WGA+ vesicles) and a second fraction which failed to agglutinate (WGA- vesicles). These two membrane fractions had quantitatively different rates of Na+/K+-dependent, ouabain-sensitive ATPase and Na+/Ca2+ exchange activities, yet a similar protein composition, which suggests that they were both derived from the plasma membrane. WGA- vesicles had a decreased number of [3H]quinuclidinyl benzilate-binding sites and no detectable [3H]nitrendipine-binding sites. Electron-microscopic and freeze-fracture analysis showed that the WGA+ fraction was composed of typical spherical sarcolemmal vesicles, whereas the WGA- fraction primarily contained elongated tubular structures suggestive of the T-tubule vesicles which were previously isolated from skeletal muscle. Assays of marker enzymes revealed that these fractions were neither sarcoplasmic reticulum nor plasma membrane from endothelial cells. Moreover, WGA agglutination did not result in the separation of right-side-out and inside-out vesicles. On the basis of these findings we propose that the WGA+ fraction corresponds to highly purified sarcolemma, whereas the WGA- fraction may be derived from T-tubule membranes.

scholarly journals Concanavalin A-agglutinability of membrane-skeleton-free vesicles and aged cellular remnants derived from human erythrocytes. Is the membrane skeleton required for agglutination?

1987 ◽  
Vol 241 (2) ◽  
pp. 513-520 ◽  
Author(s):  
S M Gokhale ◽  
N G Mehta
Keyword(s):  
Concanavalin A ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Ricinus Communis ◽  
Membrane Skeleton ◽  
Con A ◽  
Intact Cells ◽  
Rigid Skeleton

Vesicles and cell remnants have been obtained by aging of erythrocytes in vitro. The vesicles lacking the membrane skeletal proteins and the remnants known to possess a rigid skeleton have been used to assess the role of membrane skeletal proteins in the process of Con A (concanavalin A)-mediated agglutination of erythrocytes. Both the vesicles and the remnants were found to bind Con A at the same density as did intact cells. The vesicles, isolated from normal as well as from the Con A-agglutinable trypsin- and Pronase-treated cells, failed to agglutinate with Con A. They were, however, well agglutinated by WGA (wheat-germ agglutinin) and RCA [Ricinus communis (castor bean) agglutinin], indicating that the vesicles are not defective in agglutination. Large, cytoskeleton-free, vesicles prepared by another procedure also gave the same results. The aged remnants from trypsin- and Pronase-treated erythrocytes showed significantly decreased agglutination with Con A, but were agglutinated as well as the fresh cells by WGA and RCA. The agglutination with Con A is thus abolished when the membrane skeleton is absent, and reduced when it is rigid, suggesting that the skeleton may play an important role in the agglutination of erythrocytes by Con A.

scholarly journals Interaction of bilirubin with human erythrocyte membranes. Bilirubin binding to neuraminidase- and phospholipase-treated membranes

1987 ◽  
Vol 248 (1) ◽  
pp. 21-26 ◽  
Author(s):  
H Sato ◽  
S Aono ◽  
R Semba ◽  
S Kashiwamata
Keyword(s):  
Human Erythrocyte ◽  
Binding Sites ◽  
Membrane Surface ◽  
Membrane Vesicles ◽  
Erythrocyte Membranes ◽  
Binding Properties ◽  
Before And After ◽  
Human Erythrocyte Membranes ◽  
Bilirubin Binding ◽  
The Right

Saturable bilirubin binding to human erythrocyte membranes was measured before and after digestion with neuraminidase and phospholipases. Neuraminidase-treated erythrocyte membranes did not show any change in their binding properties, indicating that gangliosides could be excluded as candidates for saturable bilirubin-binding sites on erythrocyte membranes. Although bilirubin-binding properties of the membranes did not change after phospholipase D digestion, either, phospholipase C treatment greatly enhanced bilirubin binding. Thus it is suggested that a negatively charged phosphoric acid moiety of phospholipids on the membrane surface may play a role to prevent a large amount of bilirubin from binding to the membranes. Further saturable bilirubin binding to inside-out sealed erythrocyte membrane vesicles showed values comparable with those of the right-side-out sealed membranes, suggesting that the bilirubin-binding sites may be distributed on both outer and inner surfaces of the membranes, or may exist in the membranes where bilirubin may be accessible from either side.

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