Skip to main content
SpringerLink
Log in

Thermodynamics and Kinetics of Unfolding of Antiparallel G-Quadruplexes in Anti-Thrombin Aptamers

  • Published:
Biochemistry (Moscow) Aims and scope Submit manuscript

Abstract

The process of unfolding of G-quadruplex structure in the RE31 DNA-aptamer and in its complex with thrombin under the action of the fluorescently labeled complementary oligonucleotides of varying length with formation of double-helix structures has been studied. It has been suggested that G-quadruplex unfolding involves formation of an intermediate complex with an oligonucleotide. Thermodynamic parameters and kinetics of unfolding of the free aptamer and its complex with thrombin differ. Extension of the oligonucleotide sequence complementary to G-quadruplex by two nucleotides to cover the so-called “hinge region” had little impact on the conformational transition of G-quadruplex of the free aptamer. However, a pronounced effect has been observed for the aptamer–protein complex. Most likely these differences could be explained by the thrombin-induced conformational transition of the aptamer involving the hinge region.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
Fig. 5.
Fig. 6.

Similar content being viewed by others

Abbreviations

15TBA:

first generation DNA aptamer

FAM:

6-carboxyfluorescein

RE31:

second generation DNA aptamer

References

  1. Keefe, A. D., Pai, S., and Ellington, A. (2010) Aptamers as therapeutics, Nat. Rev. Drug Discov., 9, 537-550, https://doi.org/10.1038/nrd3141.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Tan, S. Y., Acquah, C., Sidhu, A., Ongkudon, C. M., Yon, L. S., et al. (2016) SELEX modifications and bioanalytical techniques for aptamer–target binding characterization, Crit. Rev. Anal. Chem., 46, 521-537, https://doi.org/10.1080/10408347.2016.1157014.

    Article  CAS  PubMed  Google Scholar 

  3. Bock, L. C., Griffin, L. C., Latham, J. A., Vermaas, E. H., and Toole, J. J. (1992) Selection of single-stranded DNA molecules that bind and inhibit human thrombin, Nature, 355, 564-566, https://doi.org/10.1038/355564a0.

    Article  CAS  PubMed  Google Scholar 

  4. Dolinnaya, N. G., Yuminova, A. V., Spiridonova, V. A., Arutyunyan, A. M., and Kopylov, A. M. (2012) Coexistence of G-quadruplex and duplex domains within the secondary structure of 31-mer DNA thrombin-binding aptamer, J. Biomol. Struct. Dynamics, 30, 524-531, https://doi.org/10.1080/07391102.2012.687518.

    Article  CAS  Google Scholar 

  5. Spiridonova, V. A., Novikova, T. M., Sizov, V. A., Shashkovskaya, V. S., Titaeva, E. V., et al. (2019) DNA aptamers to thrombin exosite I. structure-function relationships and antithrombotic effects, Biochemistry (Moscow), 84, 1521-1528, https://doi.org/10.1134/S0006297919120113.

    Article  CAS  Google Scholar 

  6. Russo Krauss, I., Spiridonova, V., Pica, A., Napolitano, V., and Sica, F. (2016) Different duplex/quadruplex junctions determine the properties of anti-thrombin aptamers with mixed folding, Nucleic Acids Res., 44, 983-991, https://doi.org/10.1093/nar/gkv1384.

    Article  CAS  PubMed  Google Scholar 

  7. Vorlickova, M., Kejnovska, I., Sagi, J., Renciuk, D., Bednarova, K., et al. (2012) Circular dichroism and guanine quadruplexes, Methods, 57, 64-75, https://doi.org/10.1016/j.ymeth.2012.03.011.

    Article  CAS  PubMed  Google Scholar 

  8. Olsen, C. M., and Marky, L. A. (2010) Monitoring the temperature unfolding of G-quadruplexes by UV and circular dichroism spectroscopies and calorimetry techniques, Methods Mol. Biol., 608, 147-158, https://doi.org/10.1007/978-1-59745-363-9_10.

    Article  CAS  PubMed  Google Scholar 

  9. Karsisiotis, A. I., Hessari, N. M., Novellino, E., Spada, G. P., Randazzo, A., et al. (2011) Topological characterization of nucleic acid G-quadruplexes by UV absorption and circular dichroism, Angew. Chem. Int. Ed. Engl., 50, 10645-10648, https://doi.org/10.1002/anie.201105193.

    Article  CAS  PubMed  Google Scholar 

  10. Tran, P. L., Mergny, J. L., and Alberti, P. (2011) Stability of telomeric G-quadruplexes, Nucleic Acids Res., 39, 3282-3294, https://doi.org/10.1093/nar/gkq1292.

    Article  CAS  PubMed  Google Scholar 

  11. Shek, Y. L., Noudeh, G. D., Nazari, M., Heerklotz, H., Abu-Ghazalah, R. M., et al. (2014) Folding thermodynamics of the hybrid-1 type intramolecular human telomeric G-quadruplex, Biopolymers, 101, 216-227, https://doi.org/10.1002/bip.22317.

    Article  CAS  PubMed  Google Scholar 

  12. Petraccone, L., Spink, C., Trent, J. O., Garbett, N. C., Mekmaysy, C. S., et al. (2011) Structure and stability of higher-order human telomeric quadruplexes, J. Am. Chem. Soc., 133, 20951-20961, https://doi.org/10.1021/ja209192a.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. De Rache, A., and Mergny, J. L. (2015) Assessment of selectivity of G-quadruplex ligands via an optimised FRET melting assay, Biochimie, 115, 194-202, https://doi.org/10.1016/j.biochi.2015.06.002.

    Article  CAS  PubMed  Google Scholar 

  14. Kwok, C. K., Sherlock, M. E., and Bevilacqua, P. C. (2013) Effect of loop sequence and loop length on the intrinsic fluorescence of G-quadruplexes, Biochemistry, 52, 3019-3021, https://doi.org/10.1021/bi400139e.

    Article  CAS  PubMed  Google Scholar 

  15. Mendoza, O., Gueddouda, N. M., Boule, J. B., Bourdoncle, A., and Mergny, J. L. (2015) A fluorescence-based helicase assay: application to the screening of G-quadruplex ligands, Nucleic Acids Res., 43, e71, https://doi.org/10.1093/nar/gkv193.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Kreig, A., Calvert, J., Sanoica, J., Cullum, E., Tipanna, R., et al. (2015) G-quadruplex formation in double strand DNA probed by NMM and CV fluorescence, Nucleic Acids Res., 43, 7961-7970, https://doi.org/10.1093/nar/gkv749.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Adrian, M., Heddi, B., and Phan, A. T. (2012) NMR spectroscopy of G-quadruplexes, Methods, 57, 11-24, https://doi.org/10.1016/j.ymeth.2012.05.003.

    Article  CAS  PubMed  Google Scholar 

  18. Lim, K. W., Ng, V. C., Martin-Pintado, N., Heddi, B., and Phan, A. T. (2013) Structure of the human telomere in Na+ solution: an antiparallel (2+2) G-quadruplex scaffold reveals additional diversity, Nucleic Acids Res., 41, 10556-10562, https://doi.org/10.1093/nar/gkt771.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Russo Krauss, I., Pica, A., Merlino, A., Mazzarella, L., and Sica, F. (2013) Duplex-quadruplex motifs in a peculiar structural organization cooperatively contribute to thrombin binding of a DNA aptamer, Acta Crystallogr. D Biol. Crystallogr., 69, 2403-2411, https://doi.org/10.1107/S0907444913022269.

    Article  CAS  PubMed  Google Scholar 

  20. Troisi, R., Napolitano, V., Spiridonova, V., Russo Krauss, I., and Sica, F. (2018) Several structural motifs cooperate in determining the highly effective anti-thrombin activity of NU172 aptamer, Nucleic Acids Res., 46, 12177-12185, https://doi.org/10.1093/nar/gky990.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Spiridonova, V. A., Barinova, K. V., Glinkina, K. A., Melnichuk, A. V., Gainutdynov, A. A., et al. (2015) A family of DNA aptamers with varied duplex region length that forms complexes with thrombin and prothrombin, FEBS Lett., 589, 2043-2049, https://doi.org/10.1016/j.febslet.2015.06.020.

    Article  CAS  PubMed  Google Scholar 

  22. Mendoza, O., Elezgaray, J., and Mergny, J. (2015) Kinetics of quadruplex to duplex conversion, Biochimie, 118, 225-233, https://doi.org/10.1016/j.biochi.2015.09.031.

    Article  CAS  PubMed  Google Scholar 

  23. Spiridonova, V. A., Glinkina, K. A., Gainutdinov, A. A., and Arutyunyan, A. M. (2014) Production of thrombin complexes with DNA aptamers containing G-quadruplex and different duplexes, J. Nephrol. Ther., 4, 149-154, https://doi.org/10.4172/2161-0959.1000149.

    Article  Google Scholar 

  24. Hardin, C. C., Perry, A. G., and White, K. (2000) Thermodynamic and kinetic characterization of the dissociation and assembly of quadruplex nucleic acids, Biopolymers, 56, 147-194, https://doi.org/10.1002/1097-0282(2000/2001)56:3<147::AID-BIP10011>3.0.CO;2-N.

    Article  CAS  PubMed  Google Scholar 

  25. Bončina, M., Lah, J., Prislan, I., and Vesnaver, G. (2012) Energetic basis of human telomeric DNA folding into G-quadruplex structures, J. Am. Chem. Soc., 134, 9657-9663, https://doi.org/10.1021/ja300605n.

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991, Moscow, Russia

    Vera A. Spiridonova, Yulia O. Naumova, Polina A. Nikolaeva, Tatiana M. Novikova & Galina Ya. Kolomijtseva

Authors
  1. Vera A. Spiridonova
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yulia O. Naumova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Polina A. Nikolaeva
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Tatiana M. Novikova
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Galina Ya. Kolomijtseva
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

V. A. Spiridonova – concept and supervision of the work, writing text of the paper; Yu. O. Naumova – conducting experiments on fluorescence measurements, processing of the results, preparation of the paper text and figures; P. A. Nikolaeva – conducting CD and SPR experiments; T. M. Novikova – preparation of aptamer complexes, sample pre-treatment; G. Ya. Kolomijtseva – supervision of the work, calculation of thermodynamic parameters and rate constants, writing text of the paper.

Corresponding author

Correspondence to Galina Ya. Kolomijtseva.

Ethics declarations

The authors declare no conflict of interest in financial or any other sphere. This article does not contain any studies with human participants or animals performed by any of the authors.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Spiridonova, V.A., Naumova, Y.O., Nikolaeva, P.A. et al. Thermodynamics and Kinetics of Unfolding of Antiparallel G-Quadruplexes in Anti-Thrombin Aptamers. Biochemistry Moscow 87, 1292–1300 (2022). https://doi.org/10.1134/S0006297922110086

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1134/S0006297922110086

Keywords

Search

Navigation