pHAST (pH-Driven Aptamer Switch for Thrombin) Catch-and-Release of Target Protein.
A pH-driven DNA nanomachine based on the human α-thrombin binding aptamer was designed for the specific catch-and-release of human α-thrombin at neutral and acidic pH, respectively. In neutral conditions, the thrombin aptamer component of the nanomachine is exposed and exists in the G-quadruplex conformation required to bind to the target protein. At slightly acidic pH, the polyadenine tail of the nanomachine becomes partially protonated and A+(anti)•G(syn) mispairing results in a conformational change, causing the target protein to be released. Förster resonance energy transfer (FRET) was used to monitor conformational switching over multiple pH cycles. Electrophoretic mobility shift assay (EMSA) and fluorescence anisotropy were used to show pH dependent protein binding and release by the nanomachine. This approach could be applied generally to existing G-rich aptamers to develop novel biosensors, theranostics, and nanoswitches.
Chemistry Department, Carleton University 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada.
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EM
McConnell
Chemistry Department, Carleton University 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada.
R
R
Bolzon
Chemistry Department, Carleton University 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada.
P
P
Mezin
Centre for Biologics Evaluation, Health Canada , 251 Sir Frederick Banting, Ottawa, Ontario K1A 0K9, Canada.
G
G
Frahm
Centre for Biologics Evaluation, Health Canada , 251 Sir Frederick Banting, Ottawa, Ontario K1A 0K9, Canada.
M
M
Johnston
Chemistry Department, Carleton University 1125 Colonel By Drive, Ottawa, Ontario K1S 5B6, Canada.
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MC
DeRosa
D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D015195:Drug Design; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013917:Thrombin / Q000737:chemistry / Q000378:metabolism*
D052157:Aptamers, Nucleotide; C081676:thrombin aptamer; D013917:Thrombin