期刊
ACS NANO
卷 17, 期 11, 页码 10857-10871出版社
AMER CHEMICAL SOC
DOI: 10.1021/acsnano.3c02532
关键词
protein engineering; nanosensor; nanodisc; single-molecule electrophysiology; biolayer interferometry; real-time kinetics; protein detection
Nanopores are powerful tools for single-molecule protein sensing, but their use outside the nanopore presents challenges in transducing physical interactions into electrical signatures. This limits their widespread adoption. Researchers have developed single-polypeptide-chain nanopores that can be integrated with an optical platform, accelerating their optimization process and providing insights into their performance.
Nanopores are currentlyutilized as powerful tools forsingle-moleculeprotein sensing. The reporting signal typically requires protein analytesto enter the nanopore interior, yet a class of these sensors has emergedthat allows targeted detection free in solution. This tactic eliminatesthe spatial limitation of nanopore confinement. However, probing proteinsoutside the nanopore implies numerous challenges associated with transducingthe physical interactions in the aqueous phase into a reliable electricalsignature. Hence, it necessitates extensive engineering and tediousoptimization routes. These obstacles have prevented the widespreadadoption of these sensors. Here, we provide an experimental strategyby developing and validating single-polypeptide-chain nanopores amenableto single-molecule and bulk-phase protein detection approaches. Weutilize protein engineering, as well as nanopore and nanodisc technologies,to create nanopore sensors that can be integrated with an opticalplatform in addition to traditional electrical recordings. Using theoptical modality over an ensemble of detectors accelerates these sensors'optimization process for a specific task. It also provides insightsinto how the construction of these single-molecule nanopore sensorsinfluences their performance. These outcomes form a basis for evaluatingengineered nanopores beyond the fundamental limits of the resistive-pulsetechnique.
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