Isolation of Platelet Membranes. A Review

Jj, Sixma; Pm, Lips

doi:10.1055/s-0038-1646693

Cited by 27 publications

(13 citation statements)

References 23 publications

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“…The starting material contained < 4.8 Â 10 11 platelets or 800 mg total platelet protein. Assuming that 25% of total platelet protein is present in membranes [35] and a purity of 100%, the yield was 10±14%, which is close to the yield of 16% reported by Fauvel et al [27].…”

Section: Characterization Of Plasma Membranessupporting

confidence: 83%

Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins

Willigen

Nieuwland

Nürnberg

et al. 2000

European Journal of Biochemistry

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Human platelets contain a Na 1 /H 1 exchanger (NHE) that regulates the cytosolic pH. The role of trimeric G-proteins in NHE control was investigated in plasma membrane vesicles by measuring exchange of intravesicular protons for extravesicular Na 1 . Exchange was saturable, independent of membrane potential and inhibited by ethylisopropyl amiloride (K i 0.05 mmol´L 21 ), demonstrating the involvement of NHE-1. The G-protein activators AlF 4 2 and GMP-P(NH)P reduced exchange by increasing the K m for Na 1 from 11.3^2.1 mm to 21.6^1.4 mm (AlF 4 2 ) and 19.8^1.1 mm (GMP-P(NH)P), leaving V max and the Hill coefficient unchanged. This effect was abolished by inhibitors of G i -proteins (N-ethylmaleimide, holoenzymeand A-protomer of pertussis toxin) and by an anti-Ga Ig and GDP(b)S. Activation of G i -proteins by mastoparan and its synthetic analogue Mas7 also strongly reduced NHE activity. These data show that in platelets NHE-1 is under negative control of the G i -family of trimeric G-proteins.

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Section: Characterization Of Plasma Membranessupporting

confidence: 83%

Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins

Willigen

Nieuwland

Nürnberg

et al. 2000

European Journal of Biochemistry

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“…Several methods currently in use by most laboratories include sucrose density ultracentrifugation of washed cell homogenates or sonicates (29) and centrifugal separation of membrane vesicles obtained from glycerol osmotic lysis of washed whole platelets (2). One of the major difficulties with the present methodology is the relatively high degree of contamination of the surface plasma membranes with intracellular membranes.…”

Section: Discussionmentioning

confidence: 99%

Isolation of human platelet plasma membranes with polylysine beads.

Kinoshita¹,

Nachman²,

Minick³

1979

The Journal of Cell Biology

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Human platelet plasma membranes were isolated with polylysine beads according to the technique developed by Jacobson and Branton (1977, Science [Wash. D. C.] 195:302--304). Lactoperoxidase-catalyzed surface iodination revealed that ninefold greater 125I specific activity was associated with the membranes isolated on beads than with whole platelets. Enrichment in the bead membrane preparation of the activities of membrane marker enzymes, bis(p-nitrophenyl)phosphate phosphodiesterase and Na,K-ATPase, was 8.0 and 4.4, respectively. Contamination with enzymes of other organelles, cytochrome oxidase and beta-glucuronidase, was relatively low as compared with membranes isolated by sucrose gradient centrifugation. Analysis by SDS polyacrylamide gel electrophoresis showed that a full complement of surface glycoproteins was present on the membranes isolated with polylysine beads. The polylysine bead technique is a rapid, reproducible and efficient method for the preparation of relatively pure platelet plasma membranes.

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“…For aggregation experiments, blood platelets were washed by gel filtration and aggregation of platelets was followed as described under Methods. The used inhibitor concentrations were 25 Two other nucleotide analogs were tested for their influence on ADP binding and aggregation. AMP(CH2)PP had no influence on binding nor aggregation while AMP(CH2)P had a very weak effect on binding and aggregation at low ADP concentrations.…”

Section: Table III Effects Of Atp and Analogs Of Atp And Adp On [ 14cmentioning

confidence: 99%

Binding of adenosine diphosphate to human blood platelets and to isolated blood platelet membranes

Lips¹,

Sixma²,

Schiphorst³

1980

Biochimica et Biophysica Acta (BBA) - General Subjects

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SummaryThe equilibrium binding of 14C-labeled ADP to intact washed human blood platelets and to platelet membranes was investigated. With both intact platelets and platelet membranes a similar concentration dependence curve was found. It consisted of a curvilinear part below 20 pM and a rectilinear part above this concentration. At high ADP concentrations, the rectilinear part appeared to be saturable. Because of this, two classes of saturable ADP binding sites were proposed. ADP was partly converted to ATP and AMP with intact platelets while this conversion was virtually absent in isolated platelet membranes. ADP was bound to platelet membranes with the same type of curves found for intact platelets.The ADP binding to the high affinity system, which was stimulated by calcium ions, was nearly independent of temperature and had a pH optimum at 7.8.A number of agents were investigated for inhibiting properties. Of the sulfhydryl reagents only p-chloromercuribenzene sulfonate inhibited both high and low affinity binding systems while iodoacetamide and N-ethylmaleimide were without effect. Compounds acting via cyclic AMP on platelet aggregation, such as adenosine and cyclic AMP itself, had no influence on binding. Some nucleosidediphosphates and nucleotide analogs at a concentration of 100 gM had no, or only a slight, effect on high affinity ADP binding. For some other nucleotides inhibitor constants were determined for both platelet ADP aggregation The ATP formation found with intact platelets could be attributed to a nucleosidediphosphate kinase. It was investigated in some detail. The enzyme was magnesium dependent, had a Ql0 value of 1.41, a pH optimum at 8.0, was competitively inhibited by AMP and reacted via a ping pong mechanism.All findings described in this paper indicate that platelets as well as platelet membranes bind ADP with the same characteristics and they suggest that the high affinity binding of ADP is involved in platelet aggregation induced by ADP. The results on nucleosidediphosphate kinase did not permit a firm conclusion about the role of the enzyme in induction of platelet aggregation by ADP.

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Isolation of Platelet Membranes. A Review

Cited by 27 publications

References 23 publications

Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins

Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins

Isolation of human platelet plasma membranes with polylysine beads.

Binding of adenosine diphosphate to human blood platelets and to isolated blood platelet membranes

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Isolation of Platelet Membranes. A Review

Cited by 27 publications

References 23 publications

Negative regulation of the platelet Na+/H+ exchanger by trimeric G‐proteins

Negative regulation of the platelet Na+/H+ exchanger by trimeric G‐proteins

Isolation of human platelet plasma membranes with polylysine beads.

Binding of adenosine diphosphate to human blood platelets and to isolated blood platelet membranes

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Product

Resources

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Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins

Negative regulation of the platelet Na⁺/H⁺ exchanger by trimeric G‐proteins