Small-Molecule Analysis Based on DNA Strand Displacement Using a Bacteriorhodopsin Photoelectric Transducer: Taking ATP as an Example

• Published in: Sensors (Basel, Switzerland) Switzerland), 2023-08, Vol.23 (17), p.7453
• Main Authors: Chen, Hsiu-Mei, Wang, Wen-Chang, Chen, Hong-Ren
• Format: Article
• Language: English
• Subjects: Acids; Adenosine triphosphate; Analysis; Antibodies; ATP; Bacteriorhodopsin; Biosensors; Detectors; DNA; Electrodes; Electrolytes; Graphene; Hemoglobin; Immunoassay; Microorganisms; photoelectric biosensor; Proteins; purple membrane; Quantum dots; Reynolds number; Rhodopsin; Sensors; small molecule; strand displacement
• ISSN: 1424-8220; 1424-8220
• DOI: 10.3390/s23177453

A uniformly oriented purple membrane (PM) monolayer containing photoactive bacteriorhodopsin has recently been applied as a sensitive photoelectric transducer to assay color proteins and microbes quantitatively. This study extends its application to detecting small molecules, using adenosine triphosphate (ATP) as an example. A reverse detection method is used, which employs AuNPs labeling and specific DNA strand displacement. A PM monolayer-coated electrode is first covalently conjugated with an ATP-specific nucleic acid aptamer and then hybridized with another gold nanoparticle-labeled nucleic acid strand with a sequence that is partially complementary to the ATP aptamer, in order to significantly minimize the photocurrent that is generated by the PM. The resulting ATP-sensing chip restores its photocurrent production in the presence of ATP, and the photocurrent recovers more effectively as the ATP concentration increases. Direct and single-step ATP detection is achieved in 15 min, with detection limits of 5 nM and a dynamic range of 5 nM–0.1 mM. The sensing chip exhibits high selectivity against other ATP analogs and is satisfactorily stable in storage. The ATP-sensing chip is used to assay bacterial populations and achieves a detection limit for Bacillus subtilis and Escherichia coli of 102 and 103 CFU/mL, respectively. The demonstration shows that a variety of small molecules can be simultaneously quantified using PM-based biosensors.