Monolayer graphene field-effect sensors operating in liquid have been widely deployed for detecting a range of analyte species often under equilibrium conditions. Here we report on the real-time detection of the binding kinetics of the essential human enzyme, topoisomerase I interacting with substrate molecules (DNA probes) that are immobilized electrochemically on to monolayer graphene strips. By monitoring the field-effect characteristics of the graphene biosensor in real-time during the enzyme-substrate interactions, we are able to decipher the surface binding constant for the cleavage reaction step of topoisomerase I activity in a label-free manner. Moreover, an appropriate design of the capture probes allows us to distinctly follow the cleavage step of topoisomerase I functioning in real-time down to picomolar concentrations. The presented results are promising for future rapid screening of drugs that are being evaluated for regulating enzyme activity.

Zuccaro, L., Tesauro, C., Kurkina, T., Fiorani, P., Yu, H., Knudsen, B., et al. (2015). Real-Time Label-Free Direct Electronic Monitoring of Topoisomerase Enzyme Binding Kinetics on Graphene. ACS NANO, 9(11), 11166-11176 [10.1021/acsnano.5b05709].

Real-Time Label-Free Direct Electronic Monitoring of Topoisomerase Enzyme Binding Kinetics on Graphene

ZUCCARO, LAURA;DESIDERI, ALESSANDRO;
2015-01-01

Abstract

Monolayer graphene field-effect sensors operating in liquid have been widely deployed for detecting a range of analyte species often under equilibrium conditions. Here we report on the real-time detection of the binding kinetics of the essential human enzyme, topoisomerase I interacting with substrate molecules (DNA probes) that are immobilized electrochemically on to monolayer graphene strips. By monitoring the field-effect characteristics of the graphene biosensor in real-time during the enzyme-substrate interactions, we are able to decipher the surface binding constant for the cleavage reaction step of topoisomerase I activity in a label-free manner. Moreover, an appropriate design of the capture probes allows us to distinctly follow the cleavage step of topoisomerase I functioning in real-time down to picomolar concentrations. The presented results are promising for future rapid screening of drugs that are being evaluated for regulating enzyme activity.
2015
Pubblicato
Rilevanza internazionale
Articolo
Esperti anonimi
Settore BIO/11 - BIOLOGIA MOLECOLARE
English
biosensor; enzyme activity; field effect; human topoisomerase; label-free biosensing; Base Sequence; Biocatalysis; DNA Topoisomerases, Type I; Electronics; Graphite; Humans; Kinetics; Molecular Sequence Data; Protein Binding; Computer Systems; Staining and Labeling
Zuccaro, L., Tesauro, C., Kurkina, T., Fiorani, P., Yu, H., Knudsen, B., et al. (2015). Real-Time Label-Free Direct Electronic Monitoring of Topoisomerase Enzyme Binding Kinetics on Graphene. ACS NANO, 9(11), 11166-11176 [10.1021/acsnano.5b05709].
Zuccaro, L; Tesauro, C; Kurkina, T; Fiorani, P; Yu, H; Knudsen, B; Kern, K; Desideri, A; Balasubramanian, K
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2108/185107
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