Summary
Nonstationary electric currents are described which are generated by the Na,K-pump. Flat membrane sheets 0.2–1 μm in diameter containing a high density of oriented N,K-ATPase molecules are bound to a planar lipid bilayer acting as a capacitive electrode. In the aqueous phase adjacent to the bound membrane sheets, ATP is released within milliseconds from an inactive, photolabile precursor (“caged” ATP) by an intense flash of light. After the ATP-concentration jump, transient current and voltage signals can be recorded in the external circuit corresponding to a translocation of positive charge across the pump protein from the cytoplasmic to the extracellular side. These electrical signals which can be suppressed by inhibitors of the Na,K-ATPase require the presence of Na+ but not of K+ in the aqueous medium. The intrinsic pump currentI p (t) can be evaluated from the recorded current signal, using estimated values of the circuit parameters of the compound membrane system.I p (t) exhibits a biphasic behavior with a fast rising period, followed by a slower decline towards a small quasistationary current. The time constant of the rising phase ofI p (t) is found to depend on the rate of photochemical ATP release. Further information on the microscopic orgin of the current transient can be obtained by double-flash experiments and by chymotrypsin modification of the protein. These and other experiments indicate that the observed charge-translocation is associated with early events in the normal transport cycle. After activation by ATP, the pump goes through the first steps of the cycle and then enters a long-lived state from which return to the initial state is slow.
Similar content being viewed by others
References
Abercrombie, R., De Weer, P. 1978. Electric current generated by squid giant axon: External K and internal ADP effects.Am. J. Physiol. 235:C63-C68
Akera, T. 1981. Effects of cardiac glycosides on Na+, K+-ATPase.In: Handbook of Experimental Pharmacology. Vol 56/I, pp. 287–336. K. Greef, editor. Springer, Berlin
Apell, H.-J., Borlinghaus, R., Läuger, P. 1987. Fast charge translocations associated with partial reactions of the Na,K-pump: II. Microscopic analysis of transient currents.J. Membrane Biol. 97:179–191
Apell, H.-J., Marcus, M.M. 1986. Na,K-ATPase in artificial lipid vesicles: Influence of the concentration of mono- and divalent cations on pumping rate.Biochim. Biophys. Acta 862:254–264
Apell, H.-J., Marcus, M.M., Anner, B.M., Oetliker, H., Läuger, P. 1985. Optical study of active ion transport in lipid vesicles containing reconstituted Na,K-ATPase.J. Membrane Biol. 85:49–63
Benz, R., Janko, K. 1976. Voltage-induced capacitance relaxation of lipid bilayer membranes. Effects of membrane composition.Biochim. Biophys. Acta 455:721–738
Cantley, L.C. 1981. Structure and mechanism of the (Na,K)-ATPase.Curr. Top. Bioenerg. 11:201–237
Chapman, J.B., Johnson, E.A., Kootsey, J.M. 1983. Electrical and biochemical properties of an enzyme model of the sodium pump.J. Membrane Biol. 74:139–153
Deguchi, N., Jørgensen, P.L., Maunsbach, A.B. 1977. Ultrastructure of the sodium pump. Comparison of thin sectioning, negative staining and freeze-fracture of purified, memrbane-bound (Na+,K+)-ATPase.J. Cell. Biol. 75:619–634
De Luca, M., McElroy, W.D. 1978. Purification and properties of firefly luciferase.Methods Enzymol. 57:3–15
De Weer, P. 1984. Electrogenic pumps: Theoretical and practical considerations.In: Electrogenic Transport: Fundamental Principles and Physiological Implications M.P. Blaustein and M. Lieberman, editors. pp. 1–15, Raven, New York
De Weer, P., Gadsby, D.C. Rakowski, R.F. 1987. Voltage dependence of Na/K pump-mediated22Na efflux and current in squid giant axon.J. Physiol. (London) (in press)
De Weer, P., Rakowski, R.F. 1984. Current generated by backward-running electrogenic Na pump in squid giant axons.Nature (London) 309:450–452
Drachev, L.A., Jasaitis, A.A., Kaulen, A.D., Kondrashin, A.A., Liberman, N.A., Nemecek, I.B., Ostroumov, S.A., Semenov, A.Yu., Skulachev, V.P. 1974. Direct measurement of electric current generation by cytochrome oxidase, H+-ATPase and bacteriorhodopsin.Nature (London) 249:321–324
Eisner, D.A., Lederer, W.J. 1980. Characterization of the electrogenic sodium pump in cardiac Purkinje fibres.J. Physiol. (London) 303:441–474
Ernst, A., Böhme, H., Böger, P. 1983 Phosphorylation and introgenase activity in isolated heterocytes fromAnabaena variabilis.Biochim. Biophys. Acta 723:83–90
Fahr, A., Läuger, P., Bamberg, E., 1981. Photocurrent kinetics of purple-membrane sheets bound to planar bilayer membranes.J. Membrane Biol. 60:51–62
Fendler, K., Grell, E., Haubs, M., Bamberg, E. 1985. Pump currents generated by the purified Na+, K+-ATPase from kidney on black lipid membranes.EMBO J. 4:3079–3085
Forbush, B., III. 1984. Na+ movement in a single turnover of the Na pump.Proc. Natl. Acad. Sci. USA 81:5310–5314
Forbush, B. III. 1985. Rapid ion movements in a single turnover of the Na+ pump.In: The Sodium Pump. I. Glynn and C.L. Ellory, editors. pp. 599–611. Company of Biologists, Cambridge, U.K.
Frank, P., Mises, R. von 1961. Die Differential- und Integralgleichungen der Mechanik und Physik. Vol. 2, p. 555. Dover, New York
Gadsby, D.C. 1984. The Na/K pump of cardiac cells.Annu. Rev. Biophys. Bioeng. 13:373–398
Gadsby, D.C., Kimura, J., Noma A. 1985. Voltage dependence of Na/K pump current in isolated heart cells.Nature (London) 315:63–65
Glitsch, H.G., Pusch, H., Schumacher, T., Verdonck, F. 1982. An identification of the K activated Na pump current in sheep Purkinje fibres.Pfluegers Arch. 394:256–263
Glynn, I.M. 1984. The electrogenic sodium pump.In: M.P. Blaustein and M. Lieberman, editors. pp. 33–48. Raven Press, New York
Glynn, I.M. 1985. The Na+, K+-transporting adenosine triphosphatase.In: The Enzymes of Biological Membranes. (2nd ed.) Vol. 3. pp. 35–114. A.N. Martonosi, editor. Plenum, New York
Glynn, I.M., Hara, Y., Richards, D.E. 1984. The occlusion of sodium ions within the mammalian sodium-potassium pump: Its role in sodium transport.J. Physiol. (London) 351:531–547
Glynn, I.M., Karlish, S.J.D. 1976. ATP hydrolysis associated with an uncoupled sodium flux through the sodium pump: Evidence for allosteric effects of intracellular ATP and extracellular sodium.J. Physiol. (London) 256:465–496
Goldman, Y.E., Hibberd, M.G., Trentham, D.R. 1984. Relaxation of rabbit psoas muscle fibres from rigor by photochemical generation of adenosine-5′-triphosphate.J. Physiol. (London) 354:577–604
Hansen, U.-P., Gradmann, D., Sanders, D., Slayman, C.L. 1981. Interpretation of current-voltage relationphips for “active” ion transport systems: I. Steady-state reaction-kinetic analysis of class-I mechanisms.J. Membrane Biol. 63:165–190
Hasuo, H., Koketsu, K. 1985. Potential dependency of the electrogenic Na+-pump current in bullfrog atrial muscle.Jpn. J. Physiol. 35:89–100
Hebert, H., Jørgensen, P.L., Skriver, E., Maunsbach, A.B. 1982. Crystallization patterns of membrane-bound (Na++K+)-ATPase.Biochim. Biophys. Acta 689:571–574
Herrmann, T.R., Rayfield, G.W. 1978. The electrical response to light of bacteriorhodopsin in planar membranes.Biophys. J. 21:111–125
Hong, F.T., Montal, M. 1979. Bacteriorhodopsin in model membranes. A new component of the displacement photocurrent in the microsecond time scale.Biophys. J. 25:465–472
Isenberg, G., Trautwein, W. 1974. The effect of dihydroouabain and lithium ions on the outward current in cardiac Purkinje fibers.Pfluegers Arch. 350:41–54
Jørgensen, P.L. 1974. Isolation of the (Na++K+)-ATPase.Methods Enzymol. 32:277–290
Jøgensen, P.L., 1982. Mechanism of the Na+,K+ pump. Protein structure and conformations of the purified (Na−+K+)-ATPase.Biochim. Biophys. Acta 694:27–68
Jørgensen, P.L., Collins, J.H. 1986. Tryptic and chymotryptic cleavage sites in the sequence of α-subunit of (Na++K+)-ATPase from outer medulla of mammalian kidney.Biochim. Biophys. Acta 860:570–576
Jørgensen, P.L., Petersen, J. 1985. Chymotryptic cleavage of α-subunit in E1-forms of renal (Na++K+)-ATPase: Effects on enzymatic properties, ligand binding and cation exchange.Biochim. Biophys. Acta 821:319–333
Kaplan, J.H., III, B., Forbush, Hoffman, J.F. 1978. Rapid photolytic release of adenosine-5′-triphosphate from a protected analogue: Utilization by the Na∶K pump of human red blood cell ghosts.Biochemistry 17:1929–1935
Karlish, S.J.D., Kaplan, J.H. 1985. Pre-steady-state kinetics of Na+ transport through the Na,K-pump.In: The Sodium Pump. I. Glynn and C.L. Ellory, editors, pp. 501–506. Company of Biologists, Cambridge, U.K.
Karlish, S.J.D., Yates, D.W., Glynn, I.M. 1978. Elementary steps of the (Na++K+-ATPase mechanism, studied with formycin nucleotides.Biochim. Biophys. Acta 525:230–251
Keszthelyi, L., Ormos, P. 1980. Electrical signals associated with the photocycle of bacteriorhodopsin.FEBS Lett. 109:189–193
Lafaire, A.V., Schwarz, W. 1986. Voltage dependence of the rheogenic Na+/K+-ATPase in the membrane of oocytes ofXenopus laevis.J. Membrane Biol. 91:43–51
Läuger, P., Apell, H.-J. 1986. A microscopic model for the current-voltage behaviour of the Na,K-pump.Eur. Biophys. J. 13:309–321
Läuger, P., Lesslauer, W., Marti, E., Richter, J. 1967. Electrical properties of bimolecular phospholipid membranes.Biochim. Biophys. Acta 135:20–32
Lederer, W.J., Nelson, M.T. 1984. Sodium pump stoichiometry determined by simultaneous measurements of sodium efflux and membrane current in barnacle.J. Physiol. (London) 348:665–677
Liébecq, C., Lallemand, A., Degueldre-Guillaume, M.-J. 1963. Purification partielle et propriétés de l'apyrase de la pomme de terre.Bull. Soc. Chim. Biol. 45:573–594
Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J. 1951. Protein measurement with the Folin phenol reagents.J. Biol. Chem. 193:265–275
Mårdh, S., Post, R.L. 1977. Phosphorylation from adenosine triphosphate of sodium- and potassium-activated adenosine triphosphatase.J. Biol. Chem. 252:633–638
McCray, J.A., Herbette, L., Kihara, T., Trentham, D.R. 1980. A new approach to time-resolved studies of ATP-requiring biological systems: Laserflash photolysis of caged ATP.Proc. Natl. Acad. Sci. USA 77:7237–7241
McLaughlin, S.G.A., Dilger, J.P. 1979. The transport of protons across membranes by weak acids.Physiol. Rev. 60:825–863
Meunier, J.M., Tauc, L. 1970. Participation d'une pompe métabolique auf potential de repos de neurones d'aplysie.J. Physiol. (Paris) 62:192c-193c
Nakao, M., Gadsby, D.C. 1986. Voltage dependence of Na translocation by the Na/K pump.Nature (London) 323:628–630
Nakashima, S., Takahashi, K. 1966. Post-tetanic hyperpolarization and electrogenic Na pump in stretch receptor neuron of crayfish.J. Physiol. (London) 187:105–127
Reynolds, J.A., Johnson, E.A., Tanford, C. 1985. Incorporation of membrane potential into theoretical analysis of electrogenic ion pumps.Proc. Natl. Acad. Sci. USA 82:6869–6873
Robinson, J.D., Flashner, M.S. 1979. The (Na++K+)-activated ATPase. Enzymatic and transport properties.Biochim. Biophys. Acta 549:145–176
Sachs, J.R. 1970. Sodium movements in the human red cell.J. Gen. Physiol. 56:322–341
Sandeaux, R., Seta, P., Jeminent, G., Alleaume, M., Gavach, Cl. 1978. The influence of pH on the conductance of lipid bimolecular membranes in relation to the alkaline ion transport induced by carboxylic carriers grisorixin, alborixin and monensin.Biochim. Biophys. Acta 511:499–508
Schuurmans-Stekhoven, F., Bonting, S.L. 1981. Transport adenosin-triphosphatases: Properties and function.Physiol. Rev. 61:1–76
Schwartz, A., Nagano, K., Nakao, M., Lindenmayer, G.E., Allen, J.C. 1971. The sodium- and potassium-activated adenosinetriphosphatase system.Methods Pharmacol. 1:361–388
Skou, J.C. 1975. The (Na++K+) activated enzyme system and its relationship to transport of sodium and potassium.Q. Rev. Biophys. 7:401–431
Skriver, E., Maunsbach, A.B., Jørgensen, P.L. 1981. Formation of two-dimensional crystals in pure membrane-bound Na+, K+-ATPase.FEBS Lett. 131:219–222
Trissl, H.-W. 1985. Primary electrogenic processes in bacteriorhodopsin probed by photoelectric measurements with capacitive metal electrodes.Biochim. Biophys. Acta 806:124–135
Zampighi, G., Simon, S.A., Kyte, J., Kreman, M. 1986. Onedimensional crystals of (Na++K+)-ATPase dimers.Biochim. Biophys. Acta 854:45–57
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Borlinghaus, R., Apell, H.J. & Läuger, P. Fast charge translocations associated with partial reactions of the Na,K-pump: I. Current and voltage transients after photochemical release of ATP. J. Membrain Biol. 97, 161–178 (1987). https://doi.org/10.1007/BF01869220
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF01869220