Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

A Biosensor Study of Protein Interaction with the 20S Proteasome Core Particle

  • 28 Accesses


It becomes increasingly clear that ubiquitination of cellular proteins is not an indispensable prerequisite of their degradation in proteasomes. There are a number of proteins to be eliminated which are not pre-ubiquitinated for their recognition by the regulatory subcomplex of the 26S proteasome, but which directly interact with the 20S proteasome core particle (20S proteasome). The obligatory precondition for such interaction consists in existence of disordered (hydrophobic) fragments in the target protein. In this study we have investigated the interaction of a number of multifunctional (moonlighting) proteins (glyceraldehyde-3-phosphate dehydrogenase (GAPDH), aldolase, pyruvate kinase) and neurodegeneration-related proteins (α-synuclein, myelin basic protein) with 20S proteasome immobilized on the SPR-biosensor chip and stabilized by means of a bifunctional agent dimethyl pimelimidate (in order to prevent possible dissociation of this subcomplex). Only two of the investigated proteins (aldolase and pyruvate kinase) interacted with the immobilized 20S proteasome (Kd of 8.17 × 10–7 M and 5.56 × 10–7 M, respectively). In addition to earlier detected GAPDH ubiquitination, mass spectrometric analysis of the studied proteins revealed the presence of the ubiquitin signature (Lys-ε-Gly-Gly) only in aldolase. Oxidation of aldolase and pyruvate kinase, which promotes elimination of proteins via their direct interaction with 20S proteasome, caused a 2−3-fold decrease in their Kd values as compared with this parameter obtained for the intact proteins. The results of this study provide further evidence for direct interaction of both ubiquitinated proteins (aldolase), and non-ubiquitinated proteins (pyruvate kinase) with the 20S proteasome core particle (20S proteasome). The effectiveness of this interaction is basically equal for the ubiquitinated proteins and non-ubiquitinated proteins.

This is a preview of subscription content, log in to check access.

Fig. 1.
Fig. 2.
Fig. 3.


  1. 1

    Hershko, A. and Ciechanover, A., Annu. Rev. Biochem., 1992, vol. 61, pp. 761–807.

  2. 2

    Hershko, A. and Ciechanover, A., Annu. Rev. Biochem., 1998, vol. 67, pp. 425–479.

  3. 3

    Tanaka, K., Proc. Jpn. Acad.,Ser. B, 2009, vol. 85, pp. 12–36.

  4. 4

    Vertegaal, A.C.O., Chem. Rev., 2011, vol. 111, pp. 7923−7940.

  5. 5

    Bremm, A. and Komander, D., Trends in Biochem. Sci., 2011, vol. 36, no. 7, pp. 355–363.

  6. 6

    Kravtsova-Ivantsiv, Y. and Ciechanover, A., J. Cell. Science, 2012, vol. 125, pp. 539–548.

  7. 7

    Buneeva, O.A. and Medvedev, A.E., Biomed. Khim., 2016, vol. 62, no. 5, pp. 496–509.

  8. 8

    Sánchez-Lanzas, R. and Castaño, J.G., Biomolecules, 2014, vol. 4, pp. 1140–1154.

  9. 9

    Buneeva, O.A. and Medvedev, A.E., Biomed. Khim., 2018, vol. 64, no. 2, pp. 134–148.

  10. 10

    Dachsel, J.C., Lucking, C.B., Deeg, S., et al., FEBS Lett., 2005, vol. 579, pp. 3913–3919.

  11. 11

    Alvarez-Castelao, B., Goethals, M., Vandekerckhove, J., and Castano, J.G., Biochim. Biophys. Acta, 2014, vol. 1843, pp. 352–365.

  12. 12

    Alvarez-Castelao, B., Munoz, C., Sanchez, I., Goethals, M., Vandekerckhove, J., and Castano, J.G., Biochim. Biophys. Acta, 2012, vol. 1823, pp. 524–533.

  13. 13

    David, D.C., Layfield, R., Serpell, L., Narain, Y., Goedert, M., and Spillantini, M.G.J., J. Neurochem., 2002, vol. 83, pp. 176–185.

  14. 14

    Cardozo, C. and Michaud, C., Arch. Biochem. Biophys., 2002, vol. 408, pp. 103–110.

  15. 15

    Belogurov, A., Jr., Kuzina, E., Kudriaeva, A., Kononikhin, A., Kovalchuk, S., Surina, Y., Smirnov, I., Lomakin, Y., Bacheva, A., Stepanov, A., Karpova, Y., Lyupina, Y., Kharybin, O., Melamed, D., Ponomarenko, N., Sharova, N., Nikolaev, E., and Gabibov, A., FASEB J., 2015, vol. 29, pp. 1–13.

  16. 16

    Ben-Nissan, G. and Sharon, M., Biomolecules, 2014, vol. 4, pp. 862–884.

  17. 17

    Erales, J. and Coffino, P., Biochim. Biophys. Acta, 2014, vol. 1843,pp. 216–221.

  18. 18

    Raynes, R.I., Pomatto, L.C.D., and Davies, K.J.A., Mol. Aspects Med., 2016, vol. 50, pp. 41–55.

  19. 19

    Uversky, V.N., Oldfield, C.J., and Dunker, A.K., J. Mol. Recognit., 2005, vol. 18, pp. 343–384.

  20. 20

    Serdyuk, I.N., Molecular Biology, 2007, vol. 41, no. 2, pp. 262–277.

  21. 21

    Wright, P.E. and Dyson, H.J., J. Mol. Biol., 1999, vol. 293, pp. 321–331.

  22. 22

    Jeffery, C.J., Trends Biochem. Sci., 1999, vol. 24, pp. 8–11.

  23. 23

    Jeffery, C.J., Frontiers Genetics, 2015, vol. 6, 211.

  24. 24

    Mani, M., Chen, C., Amblee, V., Liu, H., Mathur, T., Zwicke, G., Zabad, S., Patel, B., Thakkar, J., and Jeffery, C.J., Nucl. Acids Res., 2015, vol. 43, pp. D277–D282.

  25. 25

    Gupta, V. and Bamezai, R.N.K., Protein Science, 2010, vol. 19, pp. 2031–2044.

  26. 26

    Dalby, A., Dauter, Z., and Littlechild, J.A., Protein Science, 1999, vol. 8, pp. 291–297.

  27. 27

    Buneeva, O.A., Gnedenko, O.V., Kopylov, A.T., Medvedeva, M.V., Zgoda, V.G., Ivanov, A.S., and Medvedev, A.E., Biochemistry (Moscow), 2017, vol. 82, no. 9, pp. 1338–1344.

  28. 28

    Scopes, R.K. and Stoter, A., Methods Enzymol., 1982, vol. 90, pt. E, pp. 479−490.

  29. 29

    Buneeva, O.A., Gnedenko, O.V., Medvedeva, M.V., Ivanov, A.S., and Medvedev, A.E., Biomed. Khim., 2016, vol. 62, no. 2, pp. 60–163.

  30. 30

    Kastle, M., Reega, S., Rogowska-Wrzesinska, A., and Grune, T., Free Rad. Biol. Med., 2012, vol. 53, pp. 1468–1477.

Download references


LCMS/MS analysis of proteins was performed in the Center of Collective Use “Human Proteome” at the Institute of Biomedical Chemistry.


This work was performed within the framework of the Program for Basic Research of State Academies of Sciences for 2013−2020 (biosensor and proteomic analysis) and was partially supported by the Russian Foundation for Basic Research (project no. 19-015-00073a; preparation of the 20S proteasome core particle and proteins for biosensor analysis).

Author information

Correspondence to O. A. Buneeva.

Ethics declarations

This study was performed in accordance with generally accepted rules and norms for the humane treatment of experimental animals.

Additional information

Translated by A. Medvedev

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Buneeva, O.A., Gnedenko, O.V., Medvedeva, M.V. et al. A Biosensor Study of Protein Interaction with the 20S Proteasome Core Particle. Biochem. Moscow Suppl. Ser. B 13, 324–328 (2019). https://doi.org/10.1134/S1990750819040024

Download citation


  • proteasome
  • ubiquitination
  • intrinsically disordered proteins
  • multifunctional proteins
  • optical biosensor
  • pyruvate kinase
  • aldolase