Vectorial Discrimination of Small Molecules with a Macrocycle Adaptor-Protein Nanopore System and Nanocavity-Dependent, pH Gradient-Controlled Analyte Kinetics
Year: 2025
Type: article
Abstract: Owing to their intrinsic qualities, protein nanopores became game-changers in the realm of analyte sensing, as they offer an inexpensive and label-free method for sophisticated recognition at the single-molecule level. Here, we exploit the complexation capability of nonfunctionalized γ-cyclodextrin (γ-CD), coupled with its propensity to get reversibly captured inside a wild-type α-hemolysin nanopore (α-HL), and achieve a hybrid construct endowing specific sensing of selected, 5 bases-long oligonucleotides. We find that the molecular discrimination capability of the system has a vectorial-like sensitivity and is influenced by the sidedness and geometry of γ-CD. We showcase that asymmetrical pH changes across the γ-CD-α-HL hybrid and the ensuing electro-osmotic flow offer a simple yet powerful method to control γ-CD capture and residence time inside the nanopore, highlighting the capability of programmable sensing of spatially separated analytes. Unexpectedly, the electro-osmotic flow ensued via pH changes exerted a negligible effect on host (γ-CD)-guest (analyte) interactions, suggesting the complexity arising from a combination of hydrodynamic effects in a restricted environment and electrostatics screening in hydrophobic nanoconfinement. We present evidence that the asymmetric, low pH-mediated, electro-osmotic stabilization of a γ-CD molecule inside α-HL enables probing of β-lactam antibiotic azlocillin encapsulation inside γ-CD under distinct ionization states.
Source: Analytical Chemistry
Language: English
Subfield: Biomedical Engineering
Field: Engineering
Domain: Physical Sciences
Open Access status: hybrid
Estimated APC paid: $4,500
OpenAlex ID: W4408055292