Abstract
The structure of tobacco mosaic virus (TMV) virions and stacked disk aggregates of TMV coat protein (CP) in solution was analyzed by synchrotron-based small-angle X-ray scattering (SAXS) and negative contrast transmission electron microscopy (TEM). TMV CP aggregates had a unique stability but did not have helical symmetry. According to the TEM data, they were stacked disks associated into transversely striated rod-shaped structures 300 to 800 Å long. According to modeling based on the crystallographic model of the 4-layer TMV CP aggregate (PDB: 1EI7), the stacked disks represented hollow cylinders. The calculated SAXS pattern for the disks was compared to the experimental one over the entire measured range. The best correlation with the SAXS data was found for the model with the repeating central pair of discs; the SAXS curves for the stacked disks were virtually identical irrespectively of the protein isolation method. The positions of maxima on the scatter curves could be used as characteristic features of the studied samples; some of the peaks were assigned to the existing elements of the quaternary structure (periodicity of aggregate structure, virion helix pitch). Low-resolution structural data for the repolymerized TMV CP aggregates in solution under conditions similar to natural were produced for the first time. Analysis of such nano-size objects is essential for their application in biomedicine and biotechnology.
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Abbreviations
- CP:
-
coat protein
- SAXS:
-
small-angle X-ray scattering
- TEM:
-
transmission electron microscopy
- TMV:
-
tobacco mosaic virus
References
Caspar, D. L., and Namba, K. (1990) Switching in the self-assembly of tobacco mosaic virus, Adv. Biophys., 26, 157–185; doi: https://doi.org/10.1016/0065-227x(90)90011-h.
Kendall, A., McDonald, M., and Stubbs, G. (2007) Precise determination of the helical repeat of tobacco mosaic virus, Virology, 369, 226–227; doi: https://doi.org/10.1016/j.virol.2007.08.013.
Butler, P. J. (1999) Self-assembly of tobacco mosaic virus: the role of an intermediate aggregate in generating both specificity and speed, Philos. Trans. R. Soc. Lond. B Biol. Sci., 354, 537–550; doi: https://doi.org/10.1098/rstb.1999.0405.
Bloomer, A. C., Champness, J. N., Bricogne, G., Staden, R., and Klug, A. (1978) Protein disk of tobacco mosaic virus at 2.8 A resolution showing the interactions within and between subunits, Nature, 276, 362–368; doi: https://doi.org/10.1038/276362a0.
Klug, A. (1999) The tobacco mosaic virus particle: structure and assembly, Philos. Trans. R. Soc. Lond. B Biol. Sci., 354, 531–535.
Butler, P. J. (1984) The current picture of the structure and assembly of tobacco mosaic virus, J. Gen. Virol., 65 (Pt. 2), 253–279; doi: https://doi.org/10.1099/0022-1317-65-2-253
Franklin, R. E., and Commoner, B. (1955) Abnormal protein associated with tobacco mosaic virus; X-ray diffraction by an abnormal protein (B8) associated with tobacco mosaic virus, Nature, 175, 1076–1077; doi: https://doi.org/10.1038/1751076a0.
Unwin, P. N., and Klug, A. (1974) Electron microscopy of the stacked disk aggregate of tobacco mosaic virus protein. I. Three-dimensional image reconstruction, J. Mol. Biol., 87, 641–656; doi: https://doi.org/10.1016/0022-2836(74)90075-8.
Dore, I., Ruhlmann, C., Oudet, P., Cahoon, M., Caspar, D. L., and Van Regenmortel, M. H. (1990) Polarity of binding of monoclonal antibodies to tobacco mosaic virus rods and stacked disks, Virology, 176, 25–29; doi: https://doi.org/10.1016/0042-6822(90)90226-h.
Diaz-Avalos, R., and Caspar, D. L. (1998) Structure of the stacked disk aggregate of tobacco mosaic virus protein, Biophys. J., 74, 595–603; doi: https://doi.org/10.1016/S0006-3495(98)77818-X.
Diaz-Avalos, R., and Caspar, D. L. (2000) Hyperstable stacked-disk structure of tobacco mosaic virus protein: electron cryomicroscopy image reconstruction related to atomic models, J. Mol. Biol., 297, 67–72; doi: https://doi.org/10.1006/jmbi.1999.3481.
Bhyravbhatla, B., Watowich, S. J., and Caspar, D. L. (1998) Refined atomic model of the four-layer aggregate of the tobacco mosaic virus coat protein at 2.4-Aresolution, Biophys. J., 74, 604–615; doi: https://doi.org/10.1016/S0006-3495(98)77819-1.
Raghavendra, K., Adams, M. L., and Schuster, T. M. (1985) Tobacco mosaic virus protein aggregates in solution: structural comparison of 20S aggregates with those near conditions for disk crystallization, Biochemistry, 24, 3298–3304; doi: https://doi.org/10.1021/bi00334a034.
Raghavendra, K., Salunke, D. M., Caspar, D. L., and Schuster, T. M. (1986) Disk aggregates of tobacco mosaic virus protein in solution: electron microscopy observations, Biochemistry, 25, 6276–6279; doi: https://doi.org/10.1021/bi00368a066.
Ksenofontov, A. L., Dobrov, E. N., Fedorova, N. V., Arutyunyan, A. M., Golanikov, A. E., Jarvekulg, L., and Shtykova, E. V. (2018) Structure of potato virus A coat protein particles and their dissociation, Mol. Biol., 52, 913–921; doi: https://doi.org/10.1134/S0026893318060109.
Ksenofontov, A. L., Dobrov, E. N., Fedorova, N. V., Serebryakova, M. V., Prusov, A. N., Baratova, L. A., Paalme, V., Jarvekulg, L., and Shtykova, E. V. (2018) Isolated potato virus A coat protein possesses unusual properties and forms different short virus-like particles, J. Biomol. Struct. Dyn., 36, 1728–1738; doi: https://doi.org/10.1080/07391102.2017.1333457.
Goodman, R. M. (1975) Reconstitution of potato virus X in vitro. I. Properties of the dissociated protein structural sub-units, Virology, 68, 287–298; doi: https://doi.org/10.1016/0042-6822(75)90272-x.
Fraenkel-Conrat, H. (1957) Degradation of tobacco mosaic virus with acetic acid, Virology, 4, 1–4; doi: https://doi.org/10.1016/0042-6822(57)90038-7.
Ksenofontov, A. L., Kozlovskii, V. S., Kordiukova, L. V., Radiukhin, V. A., Timofeeva, A. V., and Dobrov, E. N. (2006) Determination of concentration and aggregate size in influenza virus preparations from true UV absorption spectra, Mol. Biol., 40, 152–158.
Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature, 227, 680–685; doi: https://doi.org/10.1038/227680a0.
Blanchet, C. E., Spilotros, A., Schwemmer, F., Graewert, M. A., Kikhney, A., Jeffries, C. M., Franke, D., Mark, D., Zengerle, R., Cipriani, F., Fiedler, S., Roessle, M., and Svergun, D. I. (2015) Versatile sample environments and automation for biological solution X-ray scattering experiments at the P12 beamline (PETRA III, DESY), J. Appl. Crystallogr., 48, 431–443; doi: https://doi.org/10.1107/S160057671500254X.
Konarev, P. V., Volkov, V. V., Sokolova, A. V., Koch, M. H. J., and Svergun, D. I. (2003) PRIMUS: a Windows PC-based system for small-angle scattering data analysis, J. Appl. Crystallogr., 36, 1277–1282, doi: https://doi.org/10.1107/S0021889803012779.
Konarev, P. V., Petoukhov, M. V., and Svergun, D. I. (2001) MASSHA — a graphics system for rigid-body modelling of macromolecular complexes against solution scattering data, J. Appl. Crystallogr., 34, 527–532; doi: https://doi.org/10.1107/S0021889801006100.
Svergun, D., Barberato, C., and Koch, M. H. (1995) CRYSOL — a program to evaluate X-ray solution scattering of biological macromolecules from atomic coordinates, J. Appl. Crystallogr., 28, 768–773; doi: https://doi.org/10.1107/S0021889895007047.
Durham, A. C., Finch, J. T., and Klug, A. (1971) States of aggregation of tobacco mosaic virus protein, Nat. New Biol., 229, 37–42; doi: https://doi.org/10.1038/newbio229037a0.
Sachse, C., Chen, J. Z., Coureux, P. D., Stroupe, M. E., Fandrich, M., and Grigorieff, N. (2007) High-resolution electron microscopy of helical specimens: a fresh look at tobacco mosaic virus, J. Mol. Biol., 371, 812–835; doi: https://doi.org/10.1016/j.jmb.2007.05.088.
Blanchet, C. E., and Svergun, D. I. (2013) Small-angle X-ray scattering on biological macromolecules and nanocomposites in solution, Annu. Rev. Phys. Chem., 64, 37–54; doi: https://doi.org/10.1146/annurev-physchem-040412-110132.
Shtykova, E. V., Baratova, L. A., Fedorova, N. V., Radyukhin, V. A., Ksenofontov, A. L., Volkov, V. V., Shishkov, A. V., Dolgov, A. A., Shilova, L. A., Batishchev, O. V., Jeffries, C. M., and Svergun, D. I. (2013) Structural analysis of influenza A virus matrix protein M1 and its self-assemblies at low pH, PLoS One, 8, e82431; doi: https://doi.org/10.1371/journal.pone.0082431
Costa, L., Andriatis, A., Brennich, M., Teulon, J. M., Chen, S.W., Pellequer, J. L., and Round, A. (2016) Combined small angle X-ray solution scattering with atomic force microscopy for characterizing radiation damage on biological macromolecules, BMC Struct. Biol., 16, 18; doi: https://doi.org/10.1186/s12900-016-0068-2.
Spinozzi, F., Ferrero, C., Ortore, M. G., Antolinos, A. D., and Mariani, P. (2014) GENFIT: software for the analysis of small-angle X-ray and neutron scattering data of macromolecules in solution, J. Appl. Crystallogr., 47, 1132–1139; doi: https://doi.org/10.1107/S1600576714005147.
Hiragi, Y., Inoue, H., Sano, Y., Kajiwara, K., Ueki, T., Kataoka, M., Tagawa, H., Izumi, Y., Muroga, Y., and Amemiya, Y. (1988) Temperature dependence of the structure of aggregates of tobacco mosaic virus protein at pH 7.2. Static synchrotron small-angle X-ray scattering, J. Mol. Biol., 204, 129–140; doi: https://doi.org/10.1016/0022-2836(88)90604-3.
Sano, Y., Inoue, H., and Hiragi, Y. (1999) Differences of reconstitution process between tobacco mosaic virus and cucumber green mottle mosaic virus by synchrotron small angle X-ray scattering using low-temperature quenching, J. Protein Chem., 18, 801–805; doi: https://doi.org/10.1023/a:1020689720082.
Potschka, M., Koch, M. H., Adams, M. L., and Schuster, T. M. (1988) Time-resolved solution X-ray scattering of tobacco mosaic virus coat protein: kinetics and structure of intermediates, Biochemistry, 27, 8481–8491; doi: https://doi.org/10.1021/bi00422a028.
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Russian Text © The Author(s), 2020, published in Biokhimiya, 2020, Vol. 85, No. 3, pp. 360–368.
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This work supported by the Russian Foundation for Basic Research (grant 18-04-00525a) and Ministry of Science and Higher Education of the Russian Federation (State Budget Project “Crystallography and Photonics”, SAXS experiments).
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Ksenofontov, A.L., Petoukhov, M.V., Prusov, A.N. et al. Characterization of Tobacco Mosaic Virus Virions and Repolymerized Coat Protein Aggregates in Solution by Small-Angle X-Ray Scattering. Biochemistry Moscow 85, 310–317 (2020). https://doi.org/10.1134/S0006297920030062
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DOI: https://doi.org/10.1134/S0006297920030062