Kinetic optimization of a protein-responsive aptamer beacon

Bradley Hall, Sean Cater, Matt Levy, Andrew D. Ellington

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Aptamers have been utilized as biosensors because they can be readily adapted to sensor platforms and signal transduction schemes through both rational design and selection. One highly generalizable scheme for the generation of the so-called aptamer beacons involves denaturing the aptamer with antisense oligonucleotides. For example, rational design methods have been utilized to adapt anti-thrombin aptamers to function as biosensors by hybridizing an antisense oligonucleotide containing a quencher to the aptamer containing a fluorescent label. In the presence of thrombin, the binding equilibrium is shifted, the antisense oligonucleotide dissociates, and the beacon lights up. By changing the affinity of the antisense oligonucleotide for the aptamer beacon, it has proven possible to change the extent of activation of the beacon. More importantly, modulating interactions between the antisense oligonucleotide and the aptamer strongly influences the kinetics of activation. Comparisons across multiple, designed aptamer beacons indicate that there is a strong inverse correlation between the thermodynamics of hybridization and the speed of activation, a finding that should prove to be generally useful in the design of future biosensors. By pre-organizing the thrombin-binding quadruplex within the aptamer the speed of response can be greatly increased. By integrating these various interactions, we were ultimately able to design aptamer beacons that were activated by threefold within 1 min of the addition of thrombin.

Original languageEnglish (US)
Pages (from-to)1049-1059
Number of pages11
JournalBiotechnology and Bioengineering
Volume103
Issue number6
DOIs
StatePublished - Aug 15 2009

Keywords

  • Aptamer beacon
  • Biosensor
  • Kinetics
  • Signaling aptamer
  • Thermodynamics
  • Thrombin

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

Fingerprint Dive into the research topics of 'Kinetic optimization of a protein-responsive aptamer beacon'. Together they form a unique fingerprint.

  • Cite this