Skip to main content
SpringerLink
Log in

Unraveling the effect of the aptamer complementary element on the performance of duplexed aptamers: a thermodynamic study

  • Paper in Forefront
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

Duplexed aptamers (DAs) are widespread aptasensor formats that simultaneously recognize and signal the concentration of target molecules. They are composed of an aptamer and aptamer complementary element (ACE) which consists of a short oligonucleotide that partially inhibits the aptamer sequence. Although the design principles to engineer DAs are straightforward, the tailored development of DAs for a particular target is currently based on trial and error due to limited knowledge of how the ACE sequence affects the final performance of DA biosensors. Therefore, we have established a thermodynamic model describing the influence of the ACE on the performance of DAs applied in equilibrium assays and demonstrated that this relationship can be described by the binding strength between the aptamer and ACE. To validate our theoretical findings, the model was applied to the 29-mer anti-thrombin aptamer as a case study, and an experimental relation between the aptamer-ACE binding strength and performance of DAs was established. The obtained results indicated that our proposed model could accurately describe the effect of the ACE sequence on the performance of the established DAs for thrombin detection, applied for equilibrium assays. Furthermore, to characterize the binding strength between the aptamer and ACEs evaluated in this work, a set of fitting equations was derived which enables thermodynamic characterization of DNA-based interactions through thermal denaturation experiments, thereby overcoming the limitations of current predictive software and chemical denaturation experiments. Altogether, this work encourages the development, characterization, and use of DAs in the field of biosensing.

Graphical abstract

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
Fig. 7

Similar content being viewed by others

References

  1. Ellington AD, Szostak JW. In vitro selection of RNA molecules that bind specific ligands. Nature. 1990;346:818–22. https://doi.org/10.1038/346818a0.

    Article  CAS  PubMed  Google Scholar 

  2. Tombelli S, Minunni M, Mascini M. Analytical applications of aptamers. Biosens Bioelectron. 2005;20:2424–34. https://doi.org/10.1016/j.bios.2004.11.006.

    Article  CAS  PubMed  Google Scholar 

  3. Deng B, Lin Y, Wang C, Li F, Wang Z, Zhang H, et al. Aptamer binding assays for proteins: the thrombin example-a review. Anal Chim Acta. 2014;837:1–15. https://doi.org/10.1016/j.aca.2014.04.055.

    Article  CAS  PubMed  Google Scholar 

  4. Citartan M, Gopinath SCB, Tominaga J, Tan S-C, Tang T-H. Assays for aptamer-based platforms. Biosens Bioelectron. 2012;34:1–11. https://doi.org/10.1016/j.bios.2012.01.002.

    Article  CAS  PubMed  Google Scholar 

  5. Weerathunge P, Ramanathan R, Torok VA, Hodgson K, Xu Y, Goodacre R, et al. Ultrasensitive colorimetric detection of murine norovirus using nanozyme aptasensor. Anal Chem. 2019;91:3270–6. https://doi.org/10.1021/acs.analchem.8b03300.

    Article  CAS  PubMed  Google Scholar 

  6. Wang BB, Zhao X, Chen LJ, Yang C, Yan XP. Functionalized persistent luminescence nanoparticle-based aptasensor for autofluorescence-free determination of kanamycin in food samples. Anal Chem. 2021;93:2589–95. https://doi.org/10.1021/acs.analchem.0c04648.

    Article  CAS  PubMed  Google Scholar 

  7. Mo F, Han M, Weng X, Zhang Y, Li J, Li H. Magnetic-assisted methylene blue-intercalated amplified dsDNA for polarity-switching-mode photoelectrochemical aptasensing. Anal Chem. 2021;93:1764–70. https://doi.org/10.1021/acs.analchem.0c04521.

    Article  CAS  PubMed  Google Scholar 

  8. Bezerra AB, Kurian ASN, Easley CJ. Nucleic-acid driven cooperative bioassays using probe proximity or split-probe techniques. Anal Chem. 2020;93:198–214. https://doi.org/10.1021/acs.analchem.0c04364.

    Article  CAS  PubMed  Google Scholar 

  9. Munzar JD, Ng A, Juncker D. Duplexed aptamers: history, design, theory, and application to biosensing. Chem Soc Rev. 2019;48. https://doi.org/10.1039/c8cs00880a.

  10. Feagin TA, Maganzini N, Soh HT. Strategies for creating structure-switching aptamers. ACS Sensors. 2018;3:1611–5. https://doi.org/10.1021/acssensors.8b00516.

    Article  CAS  PubMed  Google Scholar 

  11. Nutiu R, Li Y. Structure-switching signaling aptamers. JACS. 2003;125:4771–8. https://doi.org/10.1021/ja028962o.

    Article  CAS  Google Scholar 

  12. Harroun SG, Prévost-Tremblay C, Lauzon D, Desrosiers A, Wang X, Pedro L, et al. Programmable DNA switches and their applications. Nanoscale. 2018;10:4607–41. https://doi.org/10.1039/c7nr07348h.

    Article  CAS  PubMed  Google Scholar 

  13. Rangel AE, Hariri AA, Eisenstein M, Soh HT. Engineering aptamer switches for multifunctional stimulus-responsive nanosystems. Adv Mater. 2020;2003704:1–26. https://doi.org/10.1002/adma.202003704.

    Article  CAS  Google Scholar 

  14. Munzar JD, Ng A, Corrado M, Juncker D. Complementary oligonucleotides regulate induced fit ligand binding in duplexed aptamers. Chem Sci. 2017:8. https://doi.org/10.1039/c6sc03993f.

  15. Munzar JD, Ng A, Juncker D. Comprehensive profiling of the ligand binding landscapes of duplexed aptamer families reveals widespread induced fit. Nat Commun. 2018:9. https://doi.org/10.1038/s41467-017-02556-3.

  16. Porchetta A, Vallée-Bélisle A, Plaxco KW, Ricci F. Using distal-site mutations and allosteric inhibition to tune, extend, and narrow the useful dynamic range of aptamer-based sensors. J Am Chem Soc. 2012;134:20601–4. https://doi.org/10.1021/ja310585e.

    Article  CAS  PubMed  Google Scholar 

  17. Bissonnette S, Del Grosso E, Simon AJ, Plaxco KW, Ricci F, Valleé-Bélisle A. Optimizing the specificity window of biomolecular receptors using structure-switching and allostery. ACS Sensors. 2020;5:1937–42. https://doi.org/10.1021/acssensors.0c00237.

    Article  CAS  PubMed  Google Scholar 

  18. Massey M, Russ Algar W, Krull UJ. Fluorescence resonance energy transfer (FRET) for DNA biosensors: FRET pairs and Förster distances for various dye-DNA conjugates. Anal Chim Acta. 2006;568:181–9. https://doi.org/10.1016/j.aca.2005.12.050.

    Article  CAS  PubMed  Google Scholar 

  19. Zadeh JN, Steenberg CD, Bois JS, Wolfe BR, Pierce MB, Khan AR, et al. NUPACK: Analysis and design of nucleic acid systems. J Comput Chem. 2011;32:170–3. https://doi.org/10.1002/jcc.

    Article  CAS  PubMed  Google Scholar 

  20. Zuker M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 2003;31. https://doi.org/10.1093/nar/gkg595.

  21. OligoAnalyzer® program, IDT, Coralville, Iowa, USA. https://www.idtdna.com/SciTools. Accessed 8 Dec 2020

  22. Moreira BG, You Y, Behlke MA, Owczarzy R. Effects of fluorescent dyes, quenchers, and dangling ends on DNA duplex stability. Biochem Biophys Res Commun. 2005;327:473–84. https://doi.org/10.1016/j.bbrc.2004.12.035.

    Article  CAS  PubMed  Google Scholar 

  23. Marras SAE, Kramer FR, Tyagi S. Efficiencies of fluorescence resonance energy transfer and contact-mediated quenching in oligonucleotide probes. Nucleic Acids Res. 2002;30:1–8. https://doi.org/10.1093/nar/gnf121.

    Article  Google Scholar 

  24. Vallée-Bélisle A, Ricci F, Plaxco KW. Thermodynamic basis for the optimization of binding-induced biomolecular switches and structure-switching biosensors. PNAS. 2009;106:13802–7.

    Article  Google Scholar 

  25. Idili A, Ricci F, Valée-Bélisle A. Determining the folding and binding free energy of DNA-based nanodevices and nanoswitches using urea titration curves. Nucleic Acids Res. 2017;45:7571–80. https://doi.org/10.1093/nar/gkx498.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Idili A, Plaxco KW, Vallé E-Bé A, Ricci F. Thermodynamic basis for engineering high-affinity, high-specificity binding-induced DNA clamp nanoswitches. ACS Nano. 2013;7:10863–9. https://doi.org/10.1021/nn404305e.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. National Center for Biotechnology Information. PubChem Compound Summary for CID 1176, Urea. https://pubchem.ncbi.nlm.nih.gov/compound/Urea. Accessed 9 Dec 2020

  28. Mergny J-L, Lacroix L. Analysis of thermal melting curves. Oligonucleotides. 2003;13:515–37.

    Article  CAS  Google Scholar 

  29. Bevington PR, Robinson DK. Error analysis in: data reduction and error analysis for the physical sciences. 3rd ed. Kent A: Peterson; 2003.

    Google Scholar 

  30. Qu JH, Dillen A, Saeys W, Lammertyn J, Spasic D. Advancements in SPR biosensing technology: an overview of recent trends in smart layers design, multiplexing concepts, continuous monitoring and in vivo sensing. Anal Chim Acta. 2020;1104:10–27. https://doi.org/10.1016/j.aca.2019.12.067.

    Article  CAS  PubMed  Google Scholar 

  31. Peeters B, Daems D, Van Der Donck T, Delport F, Lammertyn J. Real-time FO-SPR monitoring of solid-phase DNAzyme cleavage activity for cutting-edge biosensing. ACS Appl Mater Interfaces. 2019;7:6759–68. https://doi.org/10.1021/acsami.8b18756.

    Article  CAS  Google Scholar 

  32. Peeters B, Safdar S, Daems D, Goos P, Spasic D, Lammertyn J. Solid-phase PCR-amplified DNAzyme activity for real-time FO-SPR detection of the MCR-2 gene. Anal Chem. 2020;92:10783–91. https://doi.org/10.1021/acs.analchem.0c02241.

    Article  CAS  PubMed  Google Scholar 

  33. Lu J, Spasic D, Delport F, Van Stappen T, Detrez I, Daems D, et al. A rapid immunoassay for detection of infliximab in whole blood using a fiber-optic SPR biosensor. Anal Chem acsanalchem. 2017:6b05092. https://doi.org/10.1021/acs.analchem.6b05092.

  34. Vallée-Bélisle A, Ricci F, Plaxco KW. Engineering biosensors with extended, narrowed, or arbitrarily edited dynamic range. J Am Chem Soc. 2012;134:2876–9. https://doi.org/10.1021/ja209850j.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Gauglitz G. Analytical evaluation of sensor measurements. Anal Bioanal Chem. 2018;410:5–13. https://doi.org/10.1007/s00216-017-0624-z.

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

This work has received funding from Research Foundation-Flanders (FWO SB/1SC8519N).

Author information

Author notes

  1. Devin Daems

    Present address: Department of Chemistry - AXES research group, University of Antwerp, Groenenborgerlaan 171, 2010, Antwerpen, Belgium

Authors and Affiliations

  1. Department of Biosystems - Biosensors Group, KU Leuven, Willem de Croylaan 42, Box 2428, 3001, Leuven, Belgium

    Annelies Dillen & Jeroen Lammertyn

  2. Department of Microbial and Molecular Systems - Centre for Membrane Separations, Adsorption, Catalysis and Spectroscopy for Sustainable Solutions, KU Leuven, Celestijnenlaan 200F, Box 2454, 3001, Leuven, Belgium

    Wouter Vandezande

Authors
  1. Annelies Dillen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wouter Vandezande
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Devin Daems
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jeroen Lammertyn
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jeroen Lammertyn.

Ethics declarations

Conflict of interest

The authors declare no competing interests.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

DNA sequences, methods, supporting tables and figures, and mathematical derivations of equations applied in the manuscript.

ESM 1

(PDF 998 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dillen, A., Vandezande, W., Daems, D. et al. Unraveling the effect of the aptamer complementary element on the performance of duplexed aptamers: a thermodynamic study. Anal Bioanal Chem 413, 4739–4750 (2021). https://doi.org/10.1007/s00216-021-03444-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-021-03444-y

Keywords

Search

Navigation