scholarly journals Membrane-bound ATP fuels the Na/K pump. Studies on membrane-bound glycolytic enzymes on inside-out vesicles from human red cell membranes.

1981 ◽  
Vol 78 (5) ◽  
pp. 547-568 ◽  
Author(s):  
R W Mercer ◽  
P B Dunham
Keyword(s):  
Cell Membranes ◽  
Phosphoglycerate Kinase ◽  
Glycolytic Enzymes ◽  
Red Cell ◽  
Na Transport ◽  
Red Cell Membrane ◽  
Cytoplasmic Surface ◽  
Membrane Bound ◽  
Red Cell Membranes ◽  
Inside Out

ATP stimulates Na transport into inside-out vesicles (IOVs) made from human red cell membranes; strophanthidin inhibits the ATP-stimulated transport. The substrates for glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoglycerate kinase (PGK) (glycolytic enzymes bound to the cytoplasmic surface of the red cell membrane) also stimulate Na transport into IOVs without added ATP. The elution of GAPDH from the membranes prevents the stimulation by the substrates, but not by exogenous ATP. Hexokinase plus glucose (agents that promote breakdown of ATP) prevent stimulation of Na transport by exogenous ATP but not by the substrates for GAPDH and PGK. [32P]orthophosphate is incorporated into a membrane-bound organic phosphate compound shown chromatographically to be ATP. The level of membrane-bound ATP is decreased when Na is added, and this decrease is inhibited by strophanthidin. When further synthesis of [32P]ATP is blocked by the addition of unlabeled orthophosphate, all of the membrane-bound [32P]ATP is dissipated by the addition of Na. From these observations it was concluded that membrane-bound glycolytic enzymes synthesize ATP and deposit it in a membrane-associated compartment from which it is used by the Na/K pump.

Properties of the Ca2+-activated K+ channel in one-step inside-out vesicles from human red cell membranes

Nature ◽  
1982 ◽  
Vol 296 (5859) ◽  
pp. 742-744 ◽  
Author(s):  
Virgilio L. Lew ◽  
Shmuel Muallem ◽  
Carol A. Seymour
Keyword(s):  
Cell Membranes ◽  
K Channel ◽  
Red Cell ◽  
One Step ◽  
Red Cell Membranes ◽  
Inside Out

scholarly journals Temperature Dependence of Nonelectrolyte Permeation across Red Cell Membranes

1973 ◽  
Vol 61 (6) ◽  
pp. 727-746 ◽  
Author(s):  
W. R. Galey ◽  
J. D. Owen ◽  
A. K. Solomon
Keyword(s):  
Temperature Dependence ◽  
Cell Membranes ◽  
Rate Theory ◽  
Red Cell ◽  
Membrane Diffusion ◽  
Red Cell Membrane ◽  
Polar Head ◽  
Lipid Solvent ◽  
Absolute Reaction Rate ◽  
Red Cell Membranes

The temperature dependence of permeation across human red cell membranes has been determined for a series of hydrophilic and lipophilic solutes, including urea and two methyl substituted derivatives, all the straight-chain amides from formamide through valeramide and the two isomers, isobutyramide and isovaleramide. The temperature coefficient for permeation by all the hydrophilic solutes is 12 kcal mol-1 or less, whereas that for all the lipophilic solutes is 19 kcal mol-1 or greater. This difference is consonant with the view that hydrophilic molecules cross the membrane by a path different from that taken by the lipophilic ones. The thermodynamic parameters associated with lipophile permeation have been studied in detail. ΔG is negative for adsorption of lipophilic amides onto an oil-water interface, whereas it is positive for transfer of the polar head from the aqueous medium to bulk lipid solvent. Application of absolute reaction rate theory makes it possible to make a clear distinction between diffusion across the water-red cell membrane interface and diffusion within the membrane. Diffusion coefficients and apparent activation enthalpies and entropies have been computed for each process. Transfer of the polar head from the solvent into the interface is characterized by ΔG‡ = 0 kcal mol-1 and ΔS‡ negative, whereas both of these parameters have large positive values for diffusion within the membrane. Diffusion within the membrane is similar to what is expected for diffusion through a highly associated viscous fluid.

scholarly journals Spectrin tetramer-dimer equilibrium in hereditary elliptocytosis

Blood ◽  
1982 ◽  
Vol 59 (5) ◽  
pp. 900-905 ◽  
Author(s):  
T Coetzer ◽  
S Zail
Keyword(s):  
Ionic Strength ◽  
Cell Membrane ◽  
Cell Membranes ◽  
Thermal Sensitivity ◽  
Red Cells ◽  
Red Cell ◽  
Red Cell Membrane ◽  
Hereditary Elliptocytosis ◽  
Red Cell Membranes ◽  
Low Ionic Strength

The proportion of spectrin tetramers and dimers in 4 degrees C low ionic strength extracts of red cell membranes of 9 subjects with 4 different variants of hereditary elliptocytosis (HE) and 2 subjects with hereditary spherocytosis (HS) was determined by nondenaturing gel electrophoresis. Such extracts reflect the native oligomeric state of spectrin in the red cell membrane. In two hemolytic HE variants (an unclassified adult with increased thermal sensitivity of red cells and an infant also showing increased thermal sensitivity of red cells), the proportion of dimers was increased, whereas the remaining subjects had values within the control range. Conversion of spectrin tetramers to dimers under isotonic conditions at 37 degrees C, or spectrin dimers to tetramers at 30 degrees C, resulted in a high proportion of dimers in the above two HE variants, as well as in a third variant with probable mild HE and sporadic hemolysis. The mother of the infant with elliptocytosis and increased thermal sensitivity of red cells, although hematologically normal, had an increased proportion of dimers in 4 degrees C low ionic strength extracts of her red cell membranes. These findings reflect an underlying primary or secondary abnormality of spectrin in these subjects that affects the association state of spectrin in the red cell membrane. Their exact relationship to the pathogenesis of the elliptical shape of the red cell, or to the presence of hemolysis, is at present unclear.

scholarly journals Immunochemical characterization of the human blood cell membrane glycoprotein recognized by the monoclonal antibody 12E7

1987 ◽  
Vol 247 (3) ◽  
pp. 757-764 ◽  
Author(s):  
F Latron ◽  
D Blanchard ◽  
J P Cartron
Keyword(s):  
Monoclonal Antibody ◽  
Cell Membrane ◽  
Gel Electrophoresis ◽  
Cell Membranes ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Enzyme Treatment ◽  
Red Cell ◽  
Red Cell Membrane ◽  
Red Cell Membranes

The 12E7 murine monoclonal antibody recognizes a protease-sensitive component of human red cells, platelets and lymphocytes which could not be detected on granulocytes. Scatchard analyses indicated that the 125I-labelled antibody binds to 1000, 4000 and 27,000 antigen sites on each red cell, platelet and lymphocyte respectively, with a binding constant ranging from 4 × 10(7) to 9 × 10(7) M-1. The membrane components recognized by the monoclonal antibody were characterized by immunostaining on nitrocellulose sheets. A 28 kDa sialoglycoprotein was visualized following electrophoretic transfer of the red cell and lymphocyte membrane proteins separated by SDS/polyacrylamide-gel electrophoresis. Another component of 25 kDa was also clearly identified in the lymphocyte and platelet lysates, but was barely detectable in the red cell membrane preparations. Enzyme treatment of intact platelets, as well as analysis of the membrane and cytosolic preparations from these cells, have shown that the 25 kDa component was of cytoplasmic origin. The mobility of the 28 kDa membrane component is decreased following neuraminidase treatment of intact blood cells, but these cells still react normally with the monoclonal antibody, indicating that sialic acids are not required for binding. The 28 kDa component is present on red cell membranes prepared from S-s-U-, En(a-) and Gerbich(-) individuals, demonstrating that it is a new sialoglycoprotein not derived from glycophorins A, B, C or D. The 28 kDa component was totally solubilized with 0.1% Triton X-100 from red cell membranes and behaves like the other red cell membrane sialoglycoproteins since it was extracted in the aqueous phase following chloroform/methanol/water or butanol/water partitionings. The 28 kDa component could be partially purified by h.p.l.c. gel permeation chromatography and preparative SDS/polyacrylamide-gel electrophoresis. The material finally obtained strongly inhibits the 12E7 monoclonal as well as human anti-Xga antibodies, suggesting either that the 28 kDa glycoprotein carries both antigens or that the 12E7 and Xga-active molecules copurified.

Electrokinetic behavior of inside-out vesicles from human red cell membranes

1982 ◽  
Vol 689 (2) ◽  
pp. 290-298 ◽  
Author(s):  
Wei S. Yen ◽  
Robert W. Mercer ◽  
B.R. Ware ◽  
Philip B. Dunham
Keyword(s):  
Cell Membranes ◽  
Red Cell ◽  
Red Cell Membranes ◽  
Inside Out

scholarly journals Active Calcium Ion Uptake by Inside-Out and Right Side-Out Vesicles of Red Blood Cell Membranes

1972 ◽  
Vol 59 (4) ◽  
pp. 462-475 ◽  
Author(s):  
Myra L. Weiner ◽  
Kwang Soo Lee
Keyword(s):  
Cell Membrane ◽  
Cell Membranes ◽  
Gradient Centrifugation ◽  
Membrane Vesicles ◽  
Red Cell ◽  
Red Cell Membrane ◽  
Active Uptake ◽  
Ca Uptake ◽  
Red Blood Cell Membranes ◽  
Inside Out

The relationship between active extrusion of Ca++ from red cell ghosts and active uptake of Ca++ by isolated red cell membrane fragments was investigated by studying the Ca++ uptake activities of inside-out and right side-out vesicles. Preparations A and B which had mainly inside-out and right side-out vesicles, respectively, were isolated from red cell membranes and were compared with respect to Ca++ adenosine triphosphatase (ATPase) and ATP-dependent Ca++ uptake activities. Preparation A had nearly eight times more inside-out vesicles and took up eight times more 45Ca in the presence of ATP compared to preparation B. Separation of the 45Ca-labeled membrane vesicles by density gradient centrifugation showed that the 45Ca label was localized to the inside-out vesicle fraction. In addition, the 45Ca taken up in the presence of ATP was lost during a subsequent incubation in the absence of ATP. The rate of 45Ca loss was not influenced by the presence of EGTA, but was slowed in the presence of La+8 (0.1 mM) in the efflux medium. The results presented here support the thesis that the active uptake of Ca++ by red cell membrane fragments is due to the active transport of Ca++ into inside-out vesicles.

scholarly journals Mechanism of spontaneous inside-out vesiculation of red cell membranes.

1988 ◽  
Vol 106 (6) ◽  
pp. 1893-1901 ◽  
Author(s):  
V L Lew ◽  
A Hockaday ◽  
C J Freeman ◽  
R M Bookchin
Keyword(s):  
Membrane Fusion ◽  
Cell Membranes ◽  
Membrane Surface ◽  
Cell Types ◽  
Red Cell ◽  
Membrane Lysis ◽  
Red Cell Membranes ◽  
Free Membrane ◽  
Mechanical Shearing ◽  
Inside Out

In certain conditions, human red cell membranes spontaneously form inside out vesicles within 20 min after hypotonic lysis. Study of the geometry of this process now reveals that, contrary to earlier views of vesiculation by endocytosis or by the mechanical shearing of cytoskeleton-depleted membrane, lysis generates a persistent membrane edge which spontaneously curls, cuts, and splices the membrane surface to form single or concentric vesicles. Analysis of the processes by which proteins may stabilize a free membrane edge led us to formulate a novel zip-type mechanism for membrane cutting-splicing and fusion even in the absence of free edges. Such protein-led membrane fusion represents an alternative to mechanisms of membrane fusion based on phospholipid interactions, and may prove relevant to processes of secretion, endocytosis, phagocytosis, and membrane recycling in many cell types.

Independent routes for Na transport across dog red cell membranes

Nature ◽  
1975 ◽  
Vol 256 (5518) ◽  
pp. 580-582 ◽  
Author(s):  
B. C. ELFORD
Keyword(s):  
Cell Membranes ◽  
Red Cell ◽  
Na Transport ◽  
Red Cell Membranes

Effects of p-nitrophenylphosphate on Ca2+ transport in inside-out vesicles from human red-cell membranes

1983 ◽  
Vol 734 (2) ◽  
pp. 363-367 ◽  
Author(s):  
Ariel J. Caride ◽  
Alcides F. Rega ◽  
Patricio J. Garrahan
Keyword(s):  
Cell Membranes ◽  
Red Cell ◽  
Red Cell Membranes ◽  
Inside Out
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