Molecular and Cellular Biochemistry

, Volume 141, Issue 1, pp 57–63 | Cite as

The relationship of hydrophobic tubulin with membranes in neural tissue

  • Dante M. Beltramo
  • Mariana Nuñez
  • Alejandra del C. Alonso
  • Héctor S. Barra


Brain membrane preparations contain tubulin that can be extracted with Triton X-114. After the extract is allowed to partition, 8% of the total brain tubulin is isolated as a hydrophobic compound in the detergent-rich phase. Cytosolic tubulin does not show this hydrophobic behaviour since it is recovered in the aqueous phase. Membrane tubulin can be released by 0.1 M Na2CO3 treatment at pH≥11.5 in such a way that the hydrophobic tubulin is converted into the hydrophilic form. These results suggest that tubulin exists associated with some membrane component that confers the hydrophobic behaviour to tubulin. If the tissue is homogenized in microtubule-stabilizing buffer containing Triton X-100, the hydrophobic tubulin is isolated from the microtubule fraction. This result indicates that the hydrophobic tubulin isolated from membrane preparations belongs to microtubules thatin vivo are associated to membranes. Therefore, hydrophobic tubulin (tubulin-membrane component complex) can be obtained from membranes or from microtubules depending on the conditions of brain homogenization.

Key words

hydrophobic tubulin membrane tubulin tubulin microtubules 



Tris-buffered saline


2-(N-morpholine) ethane sulfonic acid


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  1. 1.
    Bhattacharyya B, Wolff J: Membrane-bound tubulin in brain and thyroid tissue. J Biol Chem 250: 7639–7646, 1975PubMedGoogle Scholar
  2. 2.
    Walters BB, Matus AI: Tubulin in postsynaptic junctional lattice. Nature 257: 496–498, 1975PubMedGoogle Scholar
  3. 3.
    Gozes I, Littauer U: The alpha subunit of tubulin preferentially associated with brain presynaptic membrane. FEBS Lett 99: 86–90, 1979PubMedGoogle Scholar
  4. 4.
    Zisapel N, Levi M, Gozes I: Tubulin: an integral protein of mammalian synaptic vesicle membranes. J Neurochem 34: 26–32, 1980PubMedGoogle Scholar
  5. 5.
    Babitch JA: Synaptic plasma membrane tubulin may be an integral constituent. J Neurochem 37: 1394–1400, 1981PubMedGoogle Scholar
  6. 6.
    Strocchi P, Brown BA, Young JD, Bonventre JA, Gilbert JM: The chracterization of tubulin in CNS membrane fraction. J Neurochem 37: 1295–1307, 1981PubMedGoogle Scholar
  7. 7.
    de Néchaud B, Wolff A, Jeantet C, Bourre JM: Characterization of tubulin in mouse brain myelin. J Neurochem 41: 1538–1544, 1983PubMedGoogle Scholar
  8. 8.
    Hargreaves AJ, Avila J: Localization and characterization of tubulin-like proteins associated with brain mitochondria. The presence of a membrane-specific isoform. J Neurochem 45: 490–496, 1985PubMedGoogle Scholar
  9. 9.
    Regula CS, Sager PR, Berlin RD: Membrane tubulin. In: D Soifer (ed) Dynamic Aspects of microtubule Biology. Annals of the New York Academy of Sciences 466: 832–842, 1986Google Scholar
  10. 10.
    Carlin RK, Grab DJ, Siekevitz P: Postmorten accumulation of tubulin in postsynaptic density preparations. J Neurochem 38: 94–100, 1982PubMedGoogle Scholar
  11. 11.
    Rodriguez JA, Barra HS: Tubulin and tubulin-colchicine complex bind to brain microsomal membrane in vitro: Mol Cel Biochem 56: 49–53, 1993Google Scholar
  12. 12.
    Beltramo DM, Alonso A del C, Barra HS: Tyrosinated, detyrosinated and acetylated tubulin isotypes in rat brain membranes. Their proportions in comparison with those in cytosol. Mol Cel Biochem 112: 173–180, 1992Google Scholar
  13. 13.
    Bordier C: Phase separation of integral membrane protein in Triton X-114 solution. J Biol Chem 256: 1604–1607, 1981PubMedGoogle Scholar
  14. 14.
    Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685, 1970PubMedGoogle Scholar
  15. 15.
    Towbin H, Staehelin T, Gordon J: Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc Nat Acad Sci 76: 4350–4354, 1979PubMedGoogle Scholar
  16. 16.
    Blose HS, Meltzer DI, Feramisco JR: 10 nm filaments are induced to collapse in living cells microinjected with monoclonal and polyclonal antibodies against tubulin. J Cell Biol 98: 847–858, 1984PubMedGoogle Scholar
  17. 17.
    Piperno G, Fuller MT: Monoclonal antibodies specific for an acetylated form of a-tubulin recognize the antigen in cilia and flagella from a variety of organisms. J Cell Biol 101: 2085–2094, 1985PubMedGoogle Scholar
  18. 18.
    Beltramo DM, Arce CA, Barra HS: Tyrosination of microtubules and nonassembled tubulin in brain slices. Eur J Biochem 162: 137–141, 1987PubMedGoogle Scholar
  19. 19.
    Sloboda RD, Rosenbaum JL: Purification and assay of microtubule-associated proteins (MAPs). Methods Enzymol 85: 409–416, 1982PubMedGoogle Scholar
  20. 20.
    Arce CA, Barra HS: Release of C-terminal tyrosina from tubulin and microtubules at steady state. Biochem J 226: 311–317, 1985PubMedGoogle Scholar
  21. 21.
    Greenwood FC, Hunter WM, Glover JS: The preparation of131I-labeled human growth hormone of high specific radioactivity. Biochem J 89: 114–123, 1963PubMedGoogle Scholar
  22. 22.
    Bradford MM: A rapid an sensitive method for the quantitation of microgram quantities of proteins utilizing the principles of dye-protein binding. Anal Biochem 72: 248–254, 1976PubMedGoogle Scholar
  23. 23.
    Fujiki Y, Hubbard AL, Fowler S, Lazarow PB: Isolation of intracellular membranes by means of sodium carbonate treatment: Application to endoplasmic reticulum. J Cell Biol 93: 97–102, 1982PubMedGoogle Scholar
  24. 24.
    Kuhn DM, Arthur RJr, Yoon H, Sankaran K: Tyrosine hydroxylase in secretory granules from bovine adrenal medulla. J Biol Chem 265: 5780–5786, 1990PubMedGoogle Scholar
  25. 25.
    Niehrs C, Stinchcombe JC, Huttner WB: Two membrane-bound forms of tyrosylprotein sultransferase as revealed by phase partitioning in Triton X-114. Eur J Cell Biol 58: 35–43, 1992PubMedGoogle Scholar
  26. 26.
    Morales M, Fifková E: Distribution of acetylated a-tubulin in brain. In situ localization and biochemical characterization. Cell Tissue Res 265: 415–423, 1991PubMedGoogle Scholar
  27. 27.
    Klausner RD, Kumar N, Weinstein JN, Blumenthal R, Flavin M: Interaction of tubulin with phospholipid vesicles. I. Association with vesicles at the phase transition. J Biol Chem 256: 5879–5885, 1981PubMedGoogle Scholar
  28. 28.
    Westrum LE, Gray EG: Microtubules associated with postsynaptic ‘thickenings’. J Neurocytol 6: 505–518, 1977PubMedGoogle Scholar
  29. 29.
    Dailey ME, Bridgman PC: Structure and organization of membrane organelles along distal microtubule segments in growth cones. J Neurosci Research 30: 242–258, 1991Google Scholar
  30. 30.
    Karecla PI, Kreis TE: Interaction of membranes of the Golgi complex with microtubulesin vitro. Eur J Cell Biol 57: 139–146, 1992PubMedGoogle Scholar
  31. 31.
    L'ernault SW, Rosenbaum JL:Clamydomonas a-tubulin is posttranslationally modified by acetylation on the e-amino group of a lysine. Biochemistry 24: 473–478, 1985PubMedGoogle Scholar
  32. 32.
    Cambray-Deakin MA, Burgoyne RD: Posttranslational modifications of a-tubulin. Acetylated and detyrosinated forms in axons of rat cerebellum. J Cell Biol 104: 1569–1574, 1987PubMedGoogle Scholar
  33. 33.
    Sale WS, Besharse JC, Piperno G: Distribution of acetylated a-tubulin in retina and inin vitro-assembled microtubules. Cell Motil Cytoskel 9: 243–253, 1988Google Scholar
  34. 34.
    Black MM, Baas PW, Humphries S: Dynamics of a-tubulin deacetylation in intact neurons. J Neurosci 9: 358–368, 1989PubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • Dante M. Beltramo
    • 1
  • Mariana Nuñez
    • 1
  • Alejandra del C. Alonso
    • 1
  • Héctor S. Barra
    • 1
  1. 1.Centro de Investigaciones en Química Biológica de Córdoba (CIQUIBIC), Facultad de Ciencias QuimicasUniversidad Nacional de CórdobaCórdobaArgentina

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