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2002 | 49 | 3 | 693-701
Article title

Functional reconstitution of Ral-binding GTPase activating protein, RLIP76, in proteoliposomes catalyzing ATP-dependent transport of glutathione conjugate of 4-hydroxynonenal.

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EN
Abstracts
EN
Earlier studies from our laboratories have shown that RLIP76, a previously described Ral-binding GTPase activating protein (Jullien-Flores et al., 1995, J. Biol. Chem. 270: 22473), is identical with the xenobiotic transporter DNP-SG ATPase, and can catalyze ATP-dependent transport of glutathione-conjugates as well as doxorubin (Awasthi et al., 2000, Biochemistry, 39: 9327). We have now reconstituted purified bacterially expressed RLIP76 in proteoliposomes, and have studied ATP-dependent uptake of the glutathione conjugate of 4-hydroxynonenal (GS-HNE) by these vesicles. Results of these studies show that RLIP76 reconstituted in proteoliposomes catalyzes ATP-dependent transport of GS-HNE against a concentration gradient. The transport of GS-HNE is saturable with respect to ATP as well as GS-HNE with Km values of 1.4 mM and 2.5 μM, respectively. These studies demonstrate that RLIP76 mediates active transport of GS-HNE, and are consistent with our previous work showing that RLIP76-mediated efflux of GS-HNE regulates the intracellular concentration of 4-HNE and thereby affects 4-HNE mediated signaling.
Publisher

Year
Volume
49
Issue
3
Pages
693-701
Physical description
Dates
published
2002
received
2002-08-27
accepted
2002-09-04
Contributors
  • Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX, U.S.A.
author
  • Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX, U.S.A.
author
  • Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX, U.S.A.
  • Department of Chemistry and Biochemistry, University of Texas at Arlington, Arlington, TX, U.S.A.
  • Department of Chemistry and Biochemistry, University of Texas at Arlington, Arlington, TX, U.S.A.
author
  • Department of Internal Medicine and Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, and Central Arkansas Veterans Healthcare System, Little Rock, AR, U.S.A
  • Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX, U.S.A.
References
  • Alin P, Danielson UH, Mannervik B. (1985) 4-Hydroxyalk-2-enals are substrates for glutathione transferase. FEBS Lett.; 179: 267-70.
  • Awasthi S, Singhal SS, Srivastava SK, Zimniak P, Bajpai KK, Saxena M, Sharma R, Ziller SA 3rd, Frenkel EP, Singh SV, He NG, Awasthi YC. (1994) Adenosine triphosphate-dependent transport of doxorubicin, daunomycin, and vinblastine in human tissues by a mechanism distinct from the P-glycoprotein. J Clin Invest.; 93: 958-65.
  • Awasthi S, Singhal SS, Srivastava SK, Torman RT, Zimniak P, Bandorowicz-Pikula J, Singh SV, Piper JT, Awasthi YC, Pikula S. (1998a) ATP-Dependent human erythrocyte glutathione-conjugate transporter. I. Purification, photoaffinity labeling, and kinetic characteristics of ATPase activity. Biochemistry.; 37: 5231-8.
  • Awasthi S, Singhal SS, Pikula S, Piper JT, Srivastava SK, Torman RT, Bandorowicz-Pikula J, Lin JT, Singh SV, Zimniak P, Awasthi YC. (1998b) ATP-Dependent human erythrocyte glutathione-conjugate transporter. II. Functional reconstitution of transport activity. Biochemistry.; 37: 5239-48.
  • Awasthi S, Cheng J, Singhal SS, Saini MK, Pandya U, Pikula S, Bandorowicz-Pikula J, Singh SV, Zimniak P, Awasthi YC. (2000) Novel function of human RLIP76: ATP-dependent transport of glutathione conjugates and doxorubicin. Biochemistry.; 39: 9327-34.
  • Awasthi S, Cheng JZ, Singhal SS, Pandya U, Sharma R, Singh SV, Zimniak P, Awasthi YC. (2001) Functional reassembly of ATP-dependent xenobiotic transport by the N- and C-terminal domains of RLIP76 and identification of ATP-binding sequences. Biochemistry.; 40: 4159-68.
  • Awasthi S, Sharma R, Singhal SS, Zimniak P, Awasthi YC. (2002) RLIP76, a novel transporter catalyzing ATP-dependent efflux of xenobiotics. Drug Metab Dispos.; (in press).
  • Bradford MM. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem.; 72: 248-54.
  • Cheng JZ, Singhal SS, Saini M, Singhal J, Piper JT, Van Kuijk FJ, Zimniak P, Awasthi YC, Awasthi S. (1999) Effects of mGST A4 transfection on 4-hydroxynonenal-mediated apoptosis and differentiation of K562 human erythroleukemia cells. Arch Biochem Biophys.; 372: 29-36.
  • Cheng JZ, Singhal SS, Sharma A, Saini M, Yang Y, Awasthi S, Zimniak P, Awasthi YC. (2001a) Transfection of mGSTA4 in HL-60 cells protects against 4-hydroxynonenal-induced apoptosis by inhibiting JNK-mediated signaling. Arch Biochem Biophys.; 392: 197-207.
  • Cheng JZ, Sharma R, Yang Y, Singhal SS, Sharma A, Saini MK, Singh SV, Zimniak P, Awasthi S, Awasthi YC. (2001b) Accelerated metabolism and exclusion of 4-hydroxynonenal through induction of RLIP76 and hGST5.8 is an early adaptive response of cells to heat and oxidative stress. J Biol Chem.; 276: 41213-23.
  • Ishikawa T. (1989) ATP/Mg2+-dependent cardiac transport system for glutathione S-conjugates. A study using rat heart sarcolemma vesicles. J Biol Chem.; 264: 17343-8.
  • Jullien-Flores V, Dorseuil O, Romero F, Letourneur F, Saragosti S, Berger R, Tavitian A, Gacon G, Camonis JH. (1995) Bridging Ral GTPase to Rho pathways. RLIP76, a Ral effector with CDC42/Rac GTPase-activating protein activity. J Biol Chem.; 270: 22473-47.
  • LaBelle EF, Singh SV, Ahmad H, Wronski L, Srivastava SK, Awasthi YC. (1988) A novel dinitrophenylglutathione-stimulated ATPase is present in human erythrocyte membranes. FEBS Lett.; 228: 53-6.
  • Renes J, de Vries EE, Hooiveld GJ, Krikken I, Jansen PL, Muller M. (2000) Multidrug resistance protein MRP1 protects against the toxicity of the major lipid peroxidation product 4-hydroxynonenal. Biochem J.; 350: 555-61.
  • Ruef J, Rao GN, Li F, Bode C, Patterson C, Bhatnagar A, Runge MS. (1998) Induction of rat aortic smooth muscle cell growth by the lipid peroxidation product 4-hydroxy-2-nonenal. Circulation.; 97: 1071-8.
  • Sharma R, Singhal SS, Cheng J, Yang Y, Sharma A, Zimniak P, Awasthi S, Awasthi YC. (2001) RLIP76 is the major ATP-dependent transporter of glutathione-conjugates and doxorubicin in human erythrocytes Arch Biochem Biophys.; 391: 171-9.
  • Singhal SS, Singhal J, Cheng J, Pikula S, Sharma R, Zimniak P, Awasthi YC, Awasthi S. (2001) Purification and functional reconstitution of intact Ral-binding GTPase activating protein, RLIP76, in artificial liposomes. Acta Biochim Polon.; 48: 551-62.
  • Srivastava S, Chandra A, Bhatnagar A, Srivastava SK, Ansari NH. (1995) Lipid peroxidation product, 4-hydroxynonenal and its conjugate with GSH, are excellent substrates of bovine lens aldose reductase. Biochem Biophys Res Commun.; 217: 741-6.
  • Srivastava S, Chandra A, Wang LF, Seifert WE Jr, DaGue BB, Ansari NH, Srivastava SK, Bhatnagar A. (1998) Metabolism of the lipid peroxidation product, 4-hydroxy- trans-2-nonenal, in isolated perfused rat heart. J Biol Chem.; 273: 10893-900.
  • Towbin H, Staehelin T, Gordon J. (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A.; 76: 4350-4.
  • Uchida K, Shiraishi M, Naito Y, Torii Y, Nakamura Y, Osawa T. (1999). Activation of stress signaling pathways by the end product of lipid peroxidation. 4-Hydroxy-2-nonenal is a potential inducer of intracellular peroxide production. J Biol Chem.; 274: 2234-42.
  • Yang Y, Cheng JZ, Singhal SS, Saini M, Pandya U, Awasthi S, Awasthi YC. (2001) Role of glutathione S-transferases in protection against lipid peroxidation. Overexpression of hGSTA2-2 in K562 cells protects against hydrogen peroxide-induced apoptosis and inhibits JNK and caspase 3 activation. J Biol Chem.; 276: 19220-30.
Document Type
Publication order reference
Identifiers
YADDA identifier
bwmeta1.element.bwnjournal-article-abpv49i3p693kz
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