Springer Protocols: Abstract: Purification of Recombinant Tau Protein and Preparation of Alzheimer-Paired Helical Filaments In Vitro

Purification of Recombinant Tau Protein and Preparation of Alzheimer-Paired Helical Filaments In Vitro

By: Stefan Barghorn², Jacek Biernat², Eckhard Mandelkow²

Abstract

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The tau protein is a neuronal microtubule-associated protein. Apart of its physiological function/3-the binding to and stabilization of microtubules/3-tau is found in Alzheimer's disease brain as insoluble fibers, the so-called “paired helical filaments” (PHFs). Investigating the fundamentals of tau polymerization is indispensable for identifying inhibitory conditions or compounds preventing PHF formation, which may slow down or even stop the degeneration of neurons in Alzheimer's disease. In this chapter, we describe the methods necessary for studying the characteristics of tau polymerization to PHFs. These include: a purification protocol for recombinantly expressed tau; a general method for the polyanion induced polymerization of tau to PHFs; the quantitation of PHFs by a fluorescence-based assay; the imaging and verification of PHFs by negative stain transmission electron microscopy.

Affiliation(s): (2) Max-Planck-Unit for Structural Molecular Biology, Hamburg, Germany

Book Title: Amyloid Proteins: Methods and Protocols

Series: Methods in Molecular Biology | Volume: 299 | Pub. Date: Dec-28-2004 | Page Range: 35-51 | DOI: 10.1385/1-59259-874-9:035

Subject: Protein Science

Key Words: Alzheimer's disease - Tau protein - paired helical filament - purification - polyanion-induced polymerization - thioflavine-S assay - electron microscopy - amyloid

References

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1.	Bond, J. P., Deverin, S. P., Inouye, H., El-Agnaf, O. M., Teeter, M. M., and Kirschner, D. A. (2003) Assemblies of Alzheimer's peptides Abeta25-35 and Abeta 31-35: reverse-turn conformation and side-chain interactions revealed by X-ray diffraction. J. Struct. Biol. 141(2), 156–170.

2.	Petkova, A. T., Ishii, Y., Balbach, J. J., et al. (2002) A structural model for Alzheimer's beta-amyloid fibrils based on experimental constraints from solid state NMR. Proc. Natl. Acad. Sci. USA 99(26), 16742–16747.

3.	Torok, M., Milton, S., Kayed, R., et al. (2002) Structural and dynamic features of Alzheimer's Abeta peptide in amyloid fibrils studied by site-directed spin labeling. J. Biol. Chem. 277(43), 40810–40815.

4.	Rochet, J. C. and Lansbury, P. T. Jr. (2000) Amyloid fibrillogenesis: themes and variations. Curr. Opin. Struct. Biol. 10(1), 60–68.

5.	Garcia, M. L. and Cleveland, D. W. (2001) Going new places using an old MAP: tau, microtubules and human neurodegenerative disease. Curr. Opin. Cell Biol. 13(1), 41–48.

6.	Goedert, M., Spillantini, M. G., and Davies, S. W. (1998) Filamentous nerve cell inclusions in neurodegenerative diseases. Curr. Opin. Neurobiol. 8(5), 619–632.

7.	Buee, L., Bussiere, T., Buee-Scherrer, V., Delacourte, A., and Hof, P. R. (2000) Tau protein isoforms, phosphorylation and role in neurodegenerative disorders. Brain Res. Rev. 33(1), 95–130.

8.	Reed, L. A., Wszolek, Z. K., and Hutton, M. (2001) Phenotypic correlations in FTDP-17. Neurobiol. Aging 22(1), 89–107.

9.	Cleveland, D. W., Hwo, S. Y., and Kirschner, M. W. (1977) Physical and chemical properties of purified tau factor and the role of tau in microtubule assembly. J. Mol. Biol. 116(2), 227–247.

10.	Lee, G., Cowan, N., and Kirschner, M. (1988) The primary structure and heterogeneity of tau protein from mouse brain. Science 239, 285–288.

11.	Schweers, O., Schonbrunn-Hanebeck, E., Marx, A., and Mandelkow, E. (1994) Structural studies of tau protein and Alzheimer paired helical filaments show no evidence for beta-structure. J. Biol. Chem. 269(39), 24290–24297.

12.	Berry, R. W., Abraha, A., Lagalwar, S., et al. (2003) Inhibition of tau polymerization by its carboxy-terminal caspase cleavage fragment. Biochemistry 42(27), 8325–8331.

13.	Minoura, K., Tomoo, K., Ishida, T., Hasegawa, H., Sasaki, M., and Taniguchi, T. (2002) Amphipathic helical behavior of the third repeat fragment in the tau microtubule-binding domain, studied by (1)H NMR spectroscopy. Biochem. Biophys. Res. Commun. 294(2), 210–214.

14.	Esposito, G., Viglino, P., Novak, M., and Cattaneo, A. (2000) The solution structure of the C-terminal segment of tau protein. J. Pept. Sci. 6(11), 550–559.

15.	Yanagawa, H., Chung, S. H., Ogawa, Y., et al. (1998) Protein anatomy: C-tail region of human tau protein as a crucial structural element in Alzheimer's paired helical filament formation in vitro. Biochemistry 37(7), 1979–1988.

16.	Kirschner, D. A., Abraham, C., and Selkoe, D. J. (1986) X-ray diffraction from intraneural paired helical filaments and extraneural amyloid fibers in Alzheimer disease indicates cross-ß conformation. Proc. Natl. Acad. Sci. USA 83, 503–507.

17.	Iqbal, K., Braak, H., Braak, E., and Grundke-Iqbal, I. (1993) Silver labeling of Alzheimer neurofibrillary changes and brain beta-amyloid. J. Histotechnology 16(N4), 335–342.

18.	von Bergen, M., Friedhoff, P., Biernat, J., Heberle, J., Mandelkow, E. M., and Mandelkow, E. (2000) Assembly of tau protein into Alzheimer paired helical filaments depends on a local sequence motif ((306)VQIVYK(311)) forming beta structure. Proc. Natl. Acad. Sci. USA 97(10), 5129–5134.

19.	vonBergen, M., Barghorn, S., Li, L., et al. (2001) Mutations of tau protein in frontotemporal dementia promote aggregation of paired helical filaments by enhancing local beta-structure. J. Biol. Chem. 276(51), 48165–48174.

20.	Giannetti, A. M., Lindwall, G., Chau, M. F., Radeke, M. J., Feinstein, S. C., and Kohlstaedt, L. A. (2000) Fibers of tau fragments, but not full length tau, exhibit a cross betastructure: implications for the formation of paired helical filaments. Protein Sci. 9(12), 2427–2435.

21.	Barghorn, S., Davies, P., and Mandelkow, E. (2004) Tau paired helical filaments from Alzheimer's disease brain and assembled in vitro are based on beta-structure in the core domain. Biochemistry 43(6), 1694–1703.

22.	Berriman, J., Serpell, L. C., Oberg, K. A., Fink, A. L., Goedert, M., and Crowther, R. A. (2003) Tau filaments from human brain and from in vitro assembly of recombinant protein show cross-beta structure. Proc. Natl. Acad. Sci. USA 100(15), 9034–9038.

23.	Perez, M., Valpuesta, J. M., Medina, M., Montejo de Garcini, E., and Avila, J. (1996) Polymerization of tau into filaments in the presence of heparin: the minimal sequence required for tau-tau interaction. J. Neurochem. 67(3), 1183–1190.

24.	Goedert, M., Jakes, R., Spillantini, M. G., Hasegawa, M., Smith, M. J., and Crowther, R. A. (1996) Assembly of microtubule-associated protein tau into Alzheimer-like filaments induced by sulphated glycosaminoglycans. Nature 383(6600), 550–553.

25.	Kampers, T., Friedhoff, P., Biernat, J., and Mandelkow, E. M. (1996) RNA stimulates aggregation of microtubule-associated protein-tau into Alzheimer-like paired helical filaments. FEBS Letters 399(3), 344–349.

26.	Wilson, D. M. and Binder, L. I. (1997) Free fatty acids stimulate the polymerization of tau and amyloid beta peptides. In vitro evidence for a common effector of pathogenesis in Alzheimer's disease. Am. J. Pathol. 150(6), 2181–2195.

27.	King, M. E., Ahuja, V., Binder, L. I., and Kuret, J. (1999) Ligand-dependent tau filament formation: implications for Alzheimer's disease progression. Biochemistry 38(45), 14851–14859.

28.	Wille, H., Drewes, G., Biernat, J., Mandelkow, E. M., and Mandelkow, E. (1992) Alzheimer-like paired helical filaments and antiparallel dimers formed from microtubule-associated protein tau in vitro. J. Cell Biol. 118, 573–584.

29.	Schweers, O., Mandelkow, E. M., Biernat, J., and Mandelkow, E. (1995) Oxidation of cysteine-322 in the repeat domain of microtubule-associated protein tau controls the in vitro assembly of paired helical filaments. Proc. Natl. Acad. Sci. USA 92(18), 8463–8467.

30.	Friedhoff, P., Schneider, A., Mandelkow, E. M., and Mandelkow, E. (1998) Rapid assembly of Alzheimer-like paired helical filaments from microtubule-associated protein tau monitored by fluorescence in solution. Biochemistry 37(28), 10223–10230.

31.	Foster, N. L., Wilhelmsen, K., Sima, A. A., Jones, M. Z., D'Amato, C. J., and Gilman, S. (1997) Frontotemporal dementia and parkinsonism linked to chromosome 17: a consensus conference. Ann. Neurol. 41(6), 706–715.

32.	Hutton, M., Lendon, C. L., Rizzu, P., et al. (1998) Association of missense and 5'-splice-site mutations in tau with the inherited dementia FTDP-17. Nature 393(6686), 702–705.

33.	Clark, L. N., Poorkaj, P., Wszolek, Z., et al. (1998) Pathogenic implications of mutations in the tau gene in pallido-ponto-nigral degeneration and related neuro-degenerative disorders linked to chromosome 17. Proc. Natl. Acad. Sci. USA 95(22), 13103–13107.

34.	Poorkaj, P., Bird, T. D., Wijsman, E., et al. (1998) Tau is a candidate gene for chromosome 17 frontotemporal dementia. Ann. Neurol. 43(6), 815–825.

35.	Goedert, M., Jakes, R., and Crowther, R. A. (1999) Effects of frontotemporal dementia FTDP-17 mutations on heparin-induced assembly of tau filaments. FEBS Lett. 450(3), 306–311.

36.	Nacharaju, P., Lewis, J., Easson, C., et al. (1999) Accelerated filament formation from tau protein with specific FTDP-17 missense mutations. FEBS Lett. 447(2–3), 195–199.

37.	Barghorn, S., Zheng-Fischhofer, Q., Ackmann, M., Biernat, J., von Bergen, M., and Mandelkow, E. (2000) Structure, microtubule interactions, and paired helical filament aggregation by tau mutants of frontotemporal dementias. Biochemistry 39(38), 11714–11721.

38.	Friedhoff, P., vonBergen, M., Mandelkow, E. M., Davies, P., and Mandelkow, E. (1998) A nucleated assembly mechanism of Alzheimer paired helical filaments. Proc. Natl. Acad. Sci. USA 95(26), 15712–15717.

39.	Wischik, C. M., Novak, M., Thogersen, H. C., et al. (1988) Isolation of a fragment of tau derived from the core of the paired helical filament of Alzheimer disease. Proc. Natl. Acad. Sci. USA 85(12), 4506–4510.

40.	Novak, M., Kabat, J., and Wischik, C. M. (1993) Molecular characterization of the minimal protease resistant tau-unit of the Alzheimer's-disease paired helical filament. EMBO J. 12, 365–370.

41.	Crowther, R. A. (1991) Straight and paired helical filaments in Alzheimer disease have a common structural unit. Proc. Natl. Acad. Sci. USA 88(6), 2288–2292.

42.	Gamblin, T. C., King, M. E., Dawson, H., et al. (2000) In vitro polymerization of tau protein monitored by laser light scattering: method and application to the study of FTDP-17 mutants. Biochemistry 39(20), 6136–6144.

43.	Kidd, M. (1963) Paired helical filaments in electron microscopy of Alzheimer's disease. Nature (Lond.) 197, 192–193.

44.	Studier, F. W., Rosenberg, A. H., Dunn, J. J., and Dubendorff, J. W. (1990) Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 185, 60–89.

45.	Biernat, J., Mandelkow, E. M., Schröter, C., et al. (1992) The switch of tau protein to an Alzheimer-like state includes the phosphorylation of two serine-proline motifs upstream of the microtubule binding region. EMBO J. 11, 1593–1597.

46.	Weingarten, M. D., Lockwood, A. H., Hwo, S. Y., and Kirschner, M. W. (1975) A protein factor essential for microtubule assembly. Proc. Natl. Acad. Sci. USA 72(5), 1858–1862.

47.	Herzog, W. and Weber, K. (1978) Fractionation of brain microtubule-associated proteins. Isolation of two different proteins which stimulate tubulin polymerization in vitro. Eur. J. Biochem. 92(1), 1–8.

48.	Gustke, N., Trinczek, B., Biernat, J., Mandelkow, E. M., and Mandelkow, E. (1994) Domains of Tau protein and interactions with microtubules. Biochemistry 33, 9511–9522.

49.	Wetlaufer, D. B. (1962) Ultraviolet spectra of proteins and amino acids. Adv. Protein Chem. 17,303–391.

50.	Saidel, L. J. and Lieberman, H. (1958) Ultraviolet absorption spectra of peptides. IV. Alanine residue. Arch. Biochem. Biophys. 76, 401–409.

51.	Goldfarb, R. (1953) Absorption spectrum of the peptide bond. II. Influence of chain length. J. Biol. Chem. 201, 317–320.

52.	Barghorn, S. and Mandelkow, E. (2002) Toward a unified scheme for the aggregation of tau into Alzheimer paired helical filaments. Biochemistry 41(50), 14885–14896.

53.	Gamblin, T. C., King, M. E., Kuret, J., Berry, R. W., and Binder, L. I. (2000) Oxidative regulation of fatty acid-induced tau polymerization. Biochemistry 39(46), 14203–14210.

54.	King, M. E., Gamblin, T. C., Kuret, J., and Binder, L. I. (2000) Differential assembly of human tau isoforms in the presence of arachidonic acid. J. Neurochem. 74(4), 1749–1757.

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