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The Use of a ?-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions.

ABSTRACT: Biosensors are becoming increasingly important and implemented in various fields such as pathogen detection, molecular diagnosis, environmental monitoring, and food safety control. In this context, we used ?-lactamases as efficient reporter enzymes in several protein-protein interaction studies. Furthermore, their ability to accept insertions of peptides or structured proteins/domains strongly encourages the use of these enzymes to generate chimeric proteins. In a recent study, we inserted a single-domain antibody fragment into the Bacillus licheniformis BlaP ?-lactamase. These small domains, also called nanobodies, are defined as the antigen-binding domains of single chain antibodies from camelids. Like common double chain antibodies, they show high affinities and specificities for their targets. The resulting chimeric protein exhibited a high affinity against its target while retaining the ?-lactamase activity. This suggests that the nanobody and ?-lactamase moieties remain functional. In the present work, we report a detailed protocol that combines our hybrid ?-lactamase system to the biosensor technology. The specific binding of the nanobody to its target can be detected thanks to a conductimetric measurement of the protons released by the catalytic activity of the enzyme.

SUBMITTER: Vandevenne M

PROVIDER: S-EPMC5912309 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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The Use of a β-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions.

Vandevenne Marylène M Dondelinger Mathieu M Yunus Sami S Freischels Astrid A Freischels Régine R Crasson Oscar O Rhazi Noureddine N Bogaerts Pierre P Galleni Moreno M Filée Patrice P

Journal of visualized experiments : JoVE 20180201 132

Biosensors are becoming increasingly important and implemented in various fields such as pathogen detection, molecular diagnosis, environmental monitoring, and food safety control. In this context, we used β-lactamases as efficient reporter enzymes in several protein-protein interaction studies. Furthermore, their ability to accept insertions of peptides or structured proteins/domains strongly encourages the use of these enzymes to generate chimeric proteins. In a recent study, we inserted a sin ...[more]

PMID: 29443069

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Dataset Information

The Use of a ?-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions.

Publications

The Use of a β-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions.

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