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Biochem J. Feb 1, 1998; 329(Pt 3): 637–646.
PMCID: PMC1219087

Interaction of phosphatidic acid and phosphatidylserine with the Ca2+-ATPase of sarcoplasmic reticulum and the mechanism of inhibition.

Abstract

The sarcoplasmic reticulum of skeletal muscle contains anionic phospholipids as well as the zwitterionic phosphatidylcholine and phosphatidylethanolamine. Here we study the effects of anionic phospholipids on the activity of the Ca2+-ATPase purified from the membrane. Reconstitution of the Ca2+-ATPase into dioleoylphosphatidylserine [di(C18:1)PS] or dioleoylphosphatidic acid [di(C18:1)PA] leads to a decrease in ATPase activity. Measurements of the quenching of the tryptophan fluorescence of the ATPase by brominated phospholipids give a relative binding constant for the anionic lipids compared with dioleoylphosphatidylcholine close to 1 and suggest that phosphatidic acid only binds to the ATPase at the bulk lipid sites around the ATPase. Addition of di(C18:1)PS or di(C18:1)PA to the ATPase in the short-chain dimyristoleoylphosphatidylcholine [di(C14:1)PC] reverse the effects of the short-chain lipid on ATPase activity and on Ca2+ binding, as revealed by the response of tryptophan fluorescence intensity to Ca2+ binding. It is concluded that the lipid headgroup and lipid fatty acyl chains have separate effects on the function of the ATPase. The anionic phospholipids have no significant effect on Ca2+ binding to the ATPase; the level of Ca2+ binding to the ATPase, the affinity of binding and the rate of dissociation of Ca2+ are unchanged by reconstitution into di(C18:1)PA. The major effect of the anionic lipids is a reduction in the maximal level of binding of MgATP. This is attributed to the formation of oligomers of the Ca2+-ATPase, in which only one molecule of the ATPase can bind MgATP dimers in di(C18:1)PS and trimers or tetramers in di(C18:1)PA. The rates of phosphorylation and dephosphorylation for the proportion of the ATPase still able to bind ATP are unaffected by reconstitution. Larger changes were observed in the level of phosphorylation of the ATPase by Pi, which became very low in the anionic phospholipids. The fluorescence response to Mg2+ for the ATPase labelled with 4-(bromomethyl)-6,7-dimethoxycoumarin was also changed in di(C18:1)PS and di(C18:1)PA, so that effects of Mg2+ became comparable with those seen on phosphorylation for the unreconstituted ATPase. The anionic phospholipids could induce a conformational change in the ATPase on binding Mg2+ equivalent to that normally induced by phosphorylation or by binding inhibitors such as thapsigargin.

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Selected References

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  • East JM, Melville D, Lee AG. Exchange rates and numbers of annular lipids for the calcium and magnesium ion dependent adenosinetriphosphatase. Biochemistry. 1985 May 21;24(11):2615–2623. [PubMed]
  • East JM, Lee AG. Lipid selectivity of the calcium and magnesium ion dependent adenosinetriphosphatase, studied with fluorescence quenching by a brominated phospholipid. Biochemistry. 1982 Aug 17;21(17):4144–4151. [PubMed]
  • London E, Feigenson GW. Fluorescence quenching in model membranes. 2. Determination of local lipid environment of the calcium adenosinetriphosphatase from sarcoplasmic reticulum. Biochemistry. 1981 Mar 31;20(7):1939–1948. [PubMed]
  • Caffrey M, Feigenson GW. Fluorescence quenching in model membranes. 3. Relationship between calcium adenosinetriphosphatase enzyme activity and the affinity of the protein for phosphatidylcholines with different acyl chain characteristics. Biochemistry. 1981 Mar 31;20(7):1949–1961. [PubMed]
  • Starling AP, East JM, Lee AG. Effects of phosphatidylcholine fatty acyl chain length on calcium binding and other functions of the (Ca(2+)-Mg2+)-ATPase. Biochemistry. 1993 Feb 16;32(6):1593–1600. [PubMed]
  • Michelangeli F, Grimes EA, East JM, Lee AG. Effects of phospholipids on the function of (Ca2(+)-Mg2+)-ATPase. Biochemistry. 1991 Jan 15;30(2):342–351. [PubMed]
  • Starling AP, East JM, Lee AG. Effects of phospholipid fatty acyl chain length on phosphorylation and dephosphorylation of the Ca(2+)-ATPase. Biochem J. 1995 Sep 15;310(Pt 3):875–879. [PMC free article] [PubMed]
  • Milting H, Heilmeyer LM, Jr, Thieleczek R. Phosphoinositides in membranes that build up the triads of rabbit skeletal muscle. FEBS Lett. 1994 May 30;345(2-3):211–218. [PubMed]
  • Enyedi A, Flura M, Sarkadi B, Gardos G, Carafoli E. The maximal velocity and the calcium affinity of the red cell calcium pump may be regulated independently. J Biol Chem. 1987 May 5;262(13):6425–6430. [PubMed]
  • Starling AP, East JM, Lee AG. Phosphatidylinositol 4-phosphate increases the rate of dephosphorylation of the phosphorylated Ca(2+)-ATPase. J Biol Chem. 1995 Jun 16;270(24):14467–14470. [PubMed]
  • Henderson IM, Khan YM, East JM, Lee AG. Binding of Ca2+ to the (Ca(2+)-Mg2+)-ATPase of sarcoplasmic reticulum: equilibrium studies. Biochem J. 1994 Feb 1;297(Pt 3):615–624. [PMC free article] [PubMed]
  • Lee AG, Baker K, Khan YM, East JM. Effects of K+ on the binding of Ca2+ to the Ca(2+)-ATPase of sarcoplasmic reticulum. Biochem J. 1995 Jan 1;305(Pt 1):225–231. [PMC free article] [PubMed]
  • Cevc G. Membrane electrostatics. Biochim Biophys Acta. 1990 Oct 8;1031(3):311–382. [PubMed]
  • Farren SB, Hope MJ, Cullis PR. Polymorphic phase preferences of phosphatidic acid: A 31P and 2H NMR study. Biochem Biophys Res Commun. 1983 Mar 16;111(2):675–682. [PubMed]
  • Papahadjopoulos D, Vail WJ, Pangborn WA, Poste G. Studies on membrane fusion. II. Induction of fusion in pure phospholipid membranes by calcium ions and other divalent metals. Biochim Biophys Acta. 1976 Oct 5;448(2):265–283. [PubMed]
  • Miner VW, Prestegard JH. Structure of divalent cation-phosphatidic acid complexes as determined by 31P-NMR. Biochim Biophys Acta. 1984 Jul 25;774(2):227–236. [PubMed]
  • Caffrey M, Feigenson GW. Influence of metal ions on the phase properties of phosphatidic acid in combination with natural and synthetic phosphatidylcholines: an X-ray diffraction study using synchrotron radiation. Biochemistry. 1984 Jan 17;23(2):323–331. [PubMed]
  • Kouaouci R, Silvius JR, Graham I, Pézolet M. Calcium-induced lateral phase separations in phosphatidylcholine-phosphatidic acid mixtures. A Raman spectroscopic study. Biochemistry. 1985 Dec 3;24(25):7132–7140. [PubMed]
  • Graham I, Gagné J, Silvius JR. Kinetics and thermodynamics of calcium-induced lateral phase separations in phosphatidic acid containing bilayers. Biochemistry. 1985 Dec 3;24(25):7123–7131. [PubMed]
  • Galla HJ, Sackmann E. Chemically induced lipid phase separation in model membranes containing charged lipids: a spin label study. Biochim Biophys Acta. 1975 Sep 2;401(3):509–529. [PubMed]
  • Ito T, Onishi S. Ca2+-induced lateral phase separations in phosphatidic acid-phosphatidylcholine membranes. Biochim Biophys Acta. 1974 May 30;352(1):29–37. [PubMed]
  • Haverstick DM, Glaser M. Visualization of Ca2+-induced phospholipid domains. Proc Natl Acad Sci U S A. 1987 Jul;84(13):4475–4479. [PMC free article] [PubMed]
  • Hope MJ, Cullis PR. Effects of divalent cations and pH on phosphatidylserine model membranes: a 31P NMR study. Biochem Biophys Res Commun. 1980 Feb 12;92(3):846–852. [PubMed]
  • de Kroon AI, Timmermans JW, Killian JA, de Kruijff B. The pH dependence of headgroup and acyl chain structure and dynamics of phosphatidylserine, studied by 2H-NMR. Chem Phys Lipids. 1990 Apr;54(1):33–42. [PubMed]
  • Browning JL, Seelig J. Bilayers of phosphatidylserine: a deuterium and phosphorus nuclear magnetic resonance study. Biochemistry. 1980 Mar 18;19(6):1262–1270. [PubMed]
  • Portis A, Newton C, Pangborn W, Papahadjopoulos D. Studies on the mechanism of membrane fusion: evidence for an intermembrane Ca2+-phospholipid complex, synergism with Mg2+, and inhibition by spectrin. Biochemistry. 1979 Mar 6;18(5):780–790. [PubMed]
  • Hauser H, Shipley GG. Interactions of divalent cations with phosphatidylserine bilayer membranes. Biochemistry. 1984 Jan 3;23(1):34–41. [PubMed]
  • Casal HL, Mantsch HH, Hauser H. Infrared and 31P-NMR studies of the interaction of Mg2+ with phosphatidylserines: effect of hydrocarbon chain unsaturation. Biochim Biophys Acta. 1989 Jul 10;982(2):228–236. [PubMed]
  • Feigenson GW. Calcium ion binding between lipid bilayers: the four-component system of phosphatidylserine, phosphatidylcholine, calcium chloride, and water. Biochemistry. 1989 Feb 7;28(3):1270–1278. [PubMed]
  • Onishi S, Ito T. Calcium-induced phase separations in phosphatidylserine--phosphatidylcholine membranes. Biochemistry. 1974 Feb 26;13(5):881–887. [PubMed]
  • Starling AP, East JM, Lee AG. Effects of gel phase phospholipid on the Ca(2+)-ATPase. Biochemistry. 1995 Mar 7;34(9):3084–3091. [PubMed]
  • Starling AP, Dalton KA, East JM, Oliver S, Lee AG. Effects of phosphatidylethanolamines on the activity of the Ca(2+)-ATPase of sarcoplasmic reticulum. Biochem J. 1996 Nov 15;320(Pt 1):309–314. [PMC free article] [PubMed]
  • Daiho T, Kubota T, Kanazawa T. Stoichiometry of tight binding of magnesium and fluoride to phosphorylation and high-affinity binding of ATP, vanadate, and calcium in the sarcoplasmic reticulum Ca(2+)-ATPase. Biochemistry. 1993 Sep 28;32(38):10021–10026. [PubMed]
  • Hardwicke PM, Green NM. The effect of delipidation on the adenosine triphosphatase of sarcoplasmic reticulum. Electron microscopy and physical properties. Eur J Biochem. 1974 Feb 15;42(1):183–193. [PubMed]
  • Stefanova HI, East JM, Gore MG, Lee AG. Labeling the (Ca(2+)-Mg2+)-ATPase of sarcoplasmic reticulum with 4-(bromomethyl)-6,7-dimethoxycoumarin: detection of conformational changes. Biochemistry. 1992 Jul 7;31(26):6023–6031. [PubMed]
  • Simmonds AC, East JM, Jones OT, Rooney EK, McWhirter J, Lee AG. Annular and non-annular binding sites on the (Ca2+ + Mg2+)-ATPase. Biochim Biophys Acta. 1982 Dec 22;693(2):398–406. [PubMed]
  • Souza DO, de Meis L. Calcium and magnesium regulation of phosphorylation by ATP and ITP in sarcoplasmic reticulum vesicles. J Biol Chem. 1976 Oct 25;251(20):6355–6359. [PubMed]
  • Starling AP, Khan YM, East JM, Lee AG. Characterization of the single Ca(2+)-binding site on the Ca(2+)-ATPase reconstituted with short- or long-chain phosphatidylcholines. Biochem J. 1994 Dec 1;304(Pt 2):569–575. [PMC free article] [PubMed]
  • Khan YM, East JM, Lee AG. Effects of pH on phosphorylation of the Ca2+-ATPase of sarcoplasmic reticulum by inorganic phosphate. Biochem J. 1997 Feb 1;321(Pt 3):671–676. [PMC free article] [PubMed]
  • Froud RJ, East JM, Rooney EK, Lee AG. Binding of long-chain alkyl derivatives to lipid bilayers and to (Ca2+-Mg2+)-ATPase. Biochemistry. 1986 Nov 18;25(23):7535–7544. [PubMed]
  • Carafoli E, Stauffer T. The plasma membrane calcium pump: functional domains, regulation of the activity, and tissue specificity of isoform expression. J Neurobiol. 1994 Mar;25(3):312–324. [PubMed]
  • Brandl CJ, Green NM, Korczak B, MacLennan DH. Two Ca2+ ATPase genes: homologies and mechanistic implications of deduced amino acid sequences. Cell. 1986 Feb 28;44(4):597–607. [PubMed]
  • Clarke DM, Loo TW, Inesi G, MacLennan DH. Location of high affinity Ca2+-binding sites within the predicted transmembrane domain of the sarcoplasmic reticulum Ca2+-ATPase. Nature. 1989 Jun 8;339(6224):476–478. [PubMed]
  • Mata AM, Stefanova HI, Gore MG, Khan YM, East JM, Lee AG. Localization of Cys-344 on the (Ca(2+)-Mg(2+)-ATPase of sarcoplasmic reticulum using resonance energy transfer. Biochim Biophys Acta. 1993 Apr 8;1147(1):6–12. [PubMed]
  • Dux L, Martonosi A. The regulation of ATPase-ATPase interactions in sarcoplasmic reticulum membrane. I. The effects of Ca2+, ATP, and inorganic phosphate. J Biol Chem. 1983 Oct 10;258(19):11896–11902. [PubMed]
  • Toyoshima C, Sasabe H, Stokes DL. Three-dimensional cryo-electron microscopy of the calcium ion pump in the sarcoplasmic reticulum membrane. Nature. 1993 Apr 1;362(6419):467–471. [PubMed]
  • Yonekura K, Stokes DL, Sasabe H, Toyoshima C. The ATP-binding site of Ca(2+)-ATPase revealed by electron image analysis. Biophys J. 1997 Mar;72(3):997–1005. [PMC free article] [PubMed]
  • Wictome M, Khan YM, East JM, Lee AG. Binding of sesquiterpene lactone inhibitors to the Ca(2+)-ATPase. Biochem J. 1995 Sep 15;310(Pt 3):859–868. [PMC free article] [PubMed]
  • Khan YM, Wictome M, East JM, Lee AG. Interactions of dihydroxybenzenes with the Ca(2+)-ATPase: separate binding sites for dihydroxybenzenes and sesquiterpene lactones. Biochemistry. 1995 Nov 7;34(44):14385–14393. [PubMed]
  • Lee AG, East JM, Jones OT, McWhirter J, Rooney EK, Simmonds AC. Binding of dansyl propranolol to the (Ca2+ + Mg2+)-ATPase. Biochim Biophys Acta. 1983 Jul 27;732(2):441–454. [PubMed]
  • Asturias FJ, Blasie JK. Location of high-affinity metal binding sites in the profile structure of the Ca+2-ATPase in the sarcoplasmic reticulum by resonance x-ray diffraction. Biophys J. 1991 Feb;59(2):488–502. [PMC free article] [PubMed]
  • Hughes G, Khan YM, East JM, Lee AG. Effects of polycations on Ca2+ binding to the Ca(2+)-ATPase. Biochem J. 1995 Jun 1;308(Pt 2):493–499. [PMC free article] [PubMed]
  • Esmann M, Marsh D. Spin-label studies on the origin of the specificity of lipid-protein interactions in Na+,K+-ATPase membranes from Squalus acanthias. Biochemistry. 1985 Jul 2;24(14):3572–3578. [PubMed]
  • Fabiato A, Fabiato F. Calculator programs for computing the composition of the solutions containing multiple metals and ligands used for experiments in skinned muscle cells. J Physiol (Paris) 1979;75(5):463–505. [PubMed]

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