Biomaterial Database

The effect of membrane potential on the mammalian sodium-potassium pump reconstituted into phospholipid vesicles. 1987

R Goldshlegger, and S J Karlish, and A Rephaeli, and W D Stein

Biochemistry Department, Weizmann Institute of Science, Rehovot, Israel.

1. We have studied effects of electrical diffusion potentials on active Na+-K+ exchange in phospholipid vesicles reconstituted with pig kidney Na+, K+-ATPase. 2. Diffusion potentials, negative inside, were established using outwardly directed K+ gradients plus valinomycin or Li+ gradients plus a Li+ ionophore, AS701. Measurement of fluorescence changes of the carbocyanine dye DiS-C3-(5) showed that the ionophores generated potentials of the expected orientation and of sufficient stability for their effects on active transport to be assessed. Measurement of rates of passive 22Na+ fluxes, over a wide range of diffusion potentials, were consistent with the quantitative predictions of the constant-field flux equation. This result demonstrates that values of diffusion potentials calculated from the Nernst or constant-field equation are accurate. 3. In some conditions, the inside-negative potential (-130 to -180 mV) accelerated the rate of ATP-dependent Na+-K+ exchange on inside-out-oriented pumps, compared to 'control' without the ionophores. Reduction in the size of the diffusion potentials by addition to the medium of Li+ with AS701 or Cs+ with the valinomycin progressively annulled the acceleratory effects, consistent with these being true effects of a change in membrane potentials. 4. At saturating cytoplasmic Na+ and ATP concentrations, the diffusion potential accelerated ATP-dependent Na+-K+ exchange by up to about 30% compared to control but this effect disappeared at rate-limiting ATP concentrations (approximately 1 microM). 5. Using prior knowledge of rate-limiting steps, we interpret this finding to mean that the conformational transition E2(2K)----E12K associated with transport of two K+ ions is voltage insensitive while E1P(3Na)----E2P3Na associated with transport of three Na+ ions is voltage sensitive. The simplest explanation is that the net charge in the transport domain of the protein when no ions, 2K+ or 3Na+ are bound is -2, 0 and +1 respectively. 6. The accelerating effect of the negative-inside diffusion potential on Na+-K+ exchange is greater at limitingly low cytoplasmic Na+ concentrations than at saturating cytoplasmic Na+ concentrations. Cytoplasmic Na+ activation curves show that the diffusion potential increases the apparent cytoplasmic Na+ affinity and reduces the sigmoidicity of cytoplasmic Na+ activation. 7. A kinetic analysis reveals that this effect on apparent affinity is due to an increase in intrinsic Na+ binding and occurs in addition to the effect on a transport rate constant.(ABSTRACT TRUNCATED AT 400 WORDS)

UI	MeSH Term	Description	Entries
D007094	Imides	Organic compounds containing two acyl groups bound to NITROGEN.	Imide
D007473	Ion Channels	Gated, ion-selective glycoproteins that traverse membranes. The stimulus for ION CHANNEL GATING can be due to a variety of stimuli such as LIGANDS, a TRANSMEMBRANE POTENTIAL DIFFERENCE, mechanical deformation or through INTRACELLULAR SIGNALING PEPTIDES AND PROTEINS.	Membrane Channels,Ion Channel,Ionic Channel,Ionic Channels,Membrane Channel,Channel, Ion,Channel, Ionic,Channel, Membrane,Channels, Ion,Channels, Ionic,Channels, Membrane
D007476	Ionophores	Chemical agents that increase the permeability of biological or artificial lipid membranes to specific ions. Most ionophores are relatively small organic molecules that act as mobile carriers within membranes or coalesce to form ion permeable channels across membranes. Many are antibiotics, and many act as uncoupling agents by short-circuiting the proton gradient across mitochondrial membranes.	Ionophore
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008322	Mammals	Warm-blooded vertebrate animals belonging to the class Mammalia, including all that possess hair and suckle their young.	Mammalia,Mammal
D008564	Membrane Potentials	The voltage differences across a membrane. For cellular membranes they are computed by subtracting the voltage measured outside the membrane from the voltage measured inside the membrane. They result from differences of inside versus outside concentration of potassium, sodium, chloride, and other ions across cells' or ORGANELLES membranes. For excitable cells, the resting membrane potentials range between -30 and -100 millivolts. Physical, chemical, or electrical stimuli can make a membrane potential more negative (hyperpolarization), or less negative (depolarization).	Resting Potentials,Transmembrane Potentials,Delta Psi,Resting Membrane Potential,Transmembrane Electrical Potential Difference,Transmembrane Potential Difference,Difference, Transmembrane Potential,Differences, Transmembrane Potential,Membrane Potential,Membrane Potential, Resting,Membrane Potentials, Resting,Potential Difference, Transmembrane,Potential Differences, Transmembrane,Potential, Membrane,Potential, Resting,Potential, Transmembrane,Potentials, Membrane,Potentials, Resting,Potentials, Transmembrane,Resting Membrane Potentials,Resting Potential,Transmembrane Potential,Transmembrane Potential Differences
D008954	Models, Biological	Theoretical representations that simulate the behavior or activity of biological processes or diseases. For disease models in living animals, DISEASE MODELS, ANIMAL is available. Biological models include the use of mathematical equations, computers, and other electronic equipment.	Biological Model,Biological Models,Model, Biological,Models, Biologic,Biologic Model,Biologic Models,Model, Biologic
D008961	Models, Structural	A representation, generally small in scale, to show the structure, construction, or appearance of something. (From Random House Unabridged Dictionary, 2d ed)	Model, Structural,Structural Model,Structural Models
D010743	Phospholipids	Lipids containing one or more phosphate groups, particularly those derived from either glycerol (phosphoglycerides see GLYCEROPHOSPHOLIPIDS) or sphingosine (SPHINGOLIPIDS). They are polar lipids that are of great importance for the structure and function of cell membranes and are the most abundant of membrane lipids, although not stored in large amounts in the system.	Phosphatides,Phospholipid
D011188	Potassium	An element in the alkali group of metals with an atomic symbol K, atomic number 19, and atomic weight 39.10. It is the chief cation in the intracellular fluid of muscle and other cells. Potassium ion is a strong electrolyte that plays a significant role in the regulation of fluid volume and maintenance of the WATER-ELECTROLYTE BALANCE.