Hydrogenated and fluorinated surfactants derived from Tris(hydroxymethyl)-acrylamidomethane allow the purification of a highly active yeast F1-F0 ATP-synthase with an enhanced stability

  • Jean-Claude Talbot
  • Alain Dautant
  • Ange Polidori
  • Bernard Pucci
  • Touria Cohen-Bouhacina
  • Abdelhamid Maali
  • Bénédicte Salin
  • Daniel Brèthes
  • Jean Velours
  • Marie-France Giraud
Article

Abstract

Loss of stability and integrity of large membrane protein complexes as well as their aggregation in a non-lipidic environment are the major bottlenecks to their structural studies. We have tested C12H25-S-poly-Tris-(hydroxymethyl)acrylamidomethane (H12-TAC) among many other detergents for extracting the yeast F1F0 ATP-synthase. H12-TAC was found to be a very efficient detergent for removing the enzyme from mitochondrial membranes without altering its sensitivity towards specific ATP-synthase inhibitors. This extracted enzyme was then solubilized by either dodecyl maltoside (DDM), H12-TAC or fluorinated surfactants such as C2H5-C6F12-C2H4-S-poly-Tris-(hydroxymethyl)acrylamidomethane (H2F6-TAC) or C6F13-C2H4-S-poly-Tris-(hydroxymethyl)acrylamidomethane (F6-TAC), two surfactants exhibiting a comparable polar head to H12-TAC but bearing a fluorinated hydrophobic tail. Preparations from enzymes purified in the presence of H12-TAC were found to be more adapted for AFM imaging than ATP-synthase purified with DDM. Keeping H12-TAC during the Ni-NTA IMAC purification step or replacing it by DDM at low concentrations did not however allow preserving enzyme activity, while fluorinated surfactants H2F6-TAC and F6-TAC were found to enhance enzyme stability and integrity as indicated by sensitivity towards inhibitors. ATPase specific activity was higher with F6-TAC than with H2F6-TAC. When enzymes were mixed with egg phosphatidylcholine, ATP-synthases purified in the presence of H2F6-TAC or F6-TAC were more stable upon time than the DDM purified enzyme. Furthermore, in the presence of lipids, an activation of ATP-synthases was observed that was transitory for enzymes purified with DDM, but lasted for weeks for ATP-synthases isolated in the presence of molecules with Tris polyalcoholic moieties. Relipidated enzymes prepared with fluorinated surfactants remained highly sensitive towards inhibitors, even after 6 weeks.

Keywords

Membrane protein complexes F1F0 ATP-synthase Detergent Fluorinated surfactants 

References

  1. Abla M, Durand G, Pucci B (2008) J. Org. Chem. 73:8142–8153CrossRefGoogle Scholar
  2. Arnold I, Pfeiffer K, Neupert W, Stuart RA, Schägger H (1998) EMBO J. 17:7170–7178CrossRefGoogle Scholar
  3. Arselin G, Giraud M-F, Dautant A, Vaillier J, Brèthes D, Coulary-Salin B, Schaeffer J, Velours J (2003) Eur. J. Biochem. 270:1875–1884Google Scholar
  4. Barthélémy P, Ameduri B, Chabaud E, Popot J-L, Pucci B (1999) Org. Lett. 1:1689–1692Google Scholar
  5. Bowie JU (2001) Curr. Opin. Struct. Biol. 11:397–402CrossRefGoogle Scholar
  6. Breyton C, Chabaud E, Chaudier Y, Pucci B, Popot J (2004) FEBS Lett. 564:312–318CrossRefGoogle Scholar
  7. Chabaud E, Barthélémy P, Mora N, Popot JL, Pucci B (1998) Biochimie. 80:515–530CrossRefGoogle Scholar
  8. Chen C, Ko Y, Delannoy M, Ludtke SJ, Chiu W, Pedersen PL (2004) J. Biol. Chem. 279:31761–31768CrossRefGoogle Scholar
  9. Dauvergne J, Polidori A, Vénien-Bryan C, Pucci B (2008) Tet. Lett. 49:2247–2250CrossRefGoogle Scholar
  10. Dickson VK, Silvester JA, Fearnley IM, Leslie AGW, Walker JE (2006) EMBO J. 25:2911–2918CrossRefGoogle Scholar
  11. Dubourg F, Aimé J-P, Marsaudon S, Couturier G, Boisgard R (2003) J. Phys. Condens. Matter. 15:6167–6177CrossRefGoogle Scholar
  12. Garavito RM, Ferguson-Miller S (2001) J. Biol. Chem. 276:32403–32406CrossRefGoogle Scholar
  13. Grandier-Vazeille X, Guérin M (1996) Anal. Biochem. 242:248–254CrossRefGoogle Scholar
  14. Guérin B, Labbe P, Somlo M (1979) Methods Enzymol. 55:149–159CrossRefGoogle Scholar
  15. Hunte C, Von Jagow G, Schagger H (2003) Membrane protein purification and crystallization. Academic, LondonGoogle Scholar
  16. Kabaleeswaran V, Puri N, Walker JE, Leslie AGW, Mueller DM (2006) EMBO J. 25:5433–5442CrossRefGoogle Scholar
  17. Kabaleeswaran V, Shen H, Symersky J, Walker J, Leslie AW, Mueller D (2009) J. Biol. Chem. 284:10546–10551CrossRefGoogle Scholar
  18. Ko YH, Delannoy M, Hullihen J, Chiu W, Pedersen PL (2003) J. Biol. Chem. 278:12305–12309CrossRefGoogle Scholar
  19. Krafft MP (2001) Adv. Drug. Deliv. Rev. 47:209–228CrossRefGoogle Scholar
  20. Lebaupain F, Salvay AG, Olivier B, Durand G, Fabiano A, Michel N, Popot J-L, Ebel C, Breyton C, Pucci B (2006) Langmuir 22:8881–8890CrossRefGoogle Scholar
  21. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) J. Biol. Chem. 193:265–275Google Scholar
  22. Lévy D, Mosser G, Lambert O, Moeck GS, Bald D, Rigaud J-L (1999) J. Struct. Biol. 127:44–52CrossRefGoogle Scholar
  23. Maali A, Hurth C, Boisgard R, Jai C, Cohen-Bouhacina T, Aimé J-P (2005) J. Appl. Phys. 97:74907.1–74907.6CrossRefGoogle Scholar
  24. MacLennan DH, Smoly JM, Tzagoloff A (1968) J. Biol. Chem. 243:1589–1597Google Scholar
  25. Myrtil E, Zarif L, Greiner J, Riess JG, Pucci B, Pavia AA (1995) J. Fluorine Chem. 71:101–105CrossRefGoogle Scholar
  26. Neff D, Tripathi S, Middendorf K, Stahlberg H, Butt H, Bamberg E, Dencher N (1997) J. Struct. Biol. 119:139–148CrossRefGoogle Scholar
  27. Park K, Berrier C, Lebaupain F, Pucci B, Popot J-L, Ghazi A, Zito F (2007) Biochem J. 403:183–187CrossRefGoogle Scholar
  28. Pavia AA, Pucci B, Riess JG, Zarif L (1991) Bioorg. Med. Chem. Lett. 1:103–106CrossRefGoogle Scholar
  29. Posokhov YO, Rodnin MV, Das SK, Pucci B, Ladokhin AS (2008) Biophys J. 95:L54–L56CrossRefGoogle Scholar
  30. Privé GG (2007) Methods. 41:388–397CrossRefGoogle Scholar
  31. Pucci B, Maurizis J, Pavia A (1991) Eur. J. Polym. 27:1101–1106CrossRefGoogle Scholar
  32. Pucci B, Maurizis J, Pavia A (1993) BioOrg. Med. Chem. Lett. 3:161–164CrossRefGoogle Scholar
  33. Rodnin MV, Posokhov YO, Contino-Pépin C, Brettmann J, Kyrychenko A, Palchevskyy SS, Pucci B, Ladokhin AS (2008) Biophys J. 94:4348–4357CrossRefGoogle Scholar
  34. Rubinstein JL, Walker JE, Henderson R (2003) EMBO J. 22:6182–6192CrossRefGoogle Scholar
  35. Schabert FA, Engel A (1994) Biophys. J. 67:2394–2403CrossRefGoogle Scholar
  36. Schägger H, Pfeiffer K (2000) EMBO J. 19:1777–1783CrossRefGoogle Scholar
  37. Schägger H, Cramer WA, von Jagow G (1994) Anal. Biochem. 217:220–230CrossRefGoogle Scholar
  38. Singleton WS, Gray MS, Brown ML, White JL (1965) J. Am. Oil. Chem. Soc. 42:53–56CrossRefGoogle Scholar
  39. Somlo M (1968) Eur. J. Biochem. 5:276–284CrossRefGoogle Scholar
  40. Stock D, Leslie AG, Walker JE (1999) Science. 286:1700–1705CrossRefGoogle Scholar
  41. Tietz A, Ochoa S (1958) Arch. Biochem. Biophys. 78:477–493CrossRefGoogle Scholar
  42. Velours J, Arselin G (2000) J. Bioenerg. Biomembr. 32:383–390CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Jean-Claude Talbot
    • 1
  • Alain Dautant
    • 1
  • Ange Polidori
    • 2
  • Bernard Pucci
    • 2
  • Touria Cohen-Bouhacina
    • 3
  • Abdelhamid Maali
    • 3
  • Bénédicte Salin
    • 1
  • Daniel Brèthes
    • 1
  • Jean Velours
    • 1
  • Marie-France Giraud
    • 1
  1. 1.CNRS, Institut de Biochimie et Génétique CellulairesUniversité Bordeaux 2Bordeaux cedexFrance
  2. 2.Faculté des Sciences, Laboratoire de Chimie Bioorganique et des systèmes moléculaires vectoriels (LCBOSMV)Université d’AvignonAvignonFrance
  3. 3.Centre de Physique Moléculaire Optique et Hertzienne (CPMOH)Université Bordeaux 1Talence cedexFrance

Personalised recommendations