Creation of a gold nanoparticle based electrochemical assay for the detection of inhibitors of bacterial cytochrome bd oxidases

• The direct electrochemical characterization of a bacterial membrane protein is shown.
• The activity of the electrodes decreases for densely packed layers of proteins.
• The electrodes can be used for identification of inhibitors the protein.

Cytochrome bd oxidases are membrane proteins expressed by bacteria including a number of pathogens, which make them an attractive target for the discovery of new antibiotics. An electrochemical assay is developed to study the activity of these proteins and inhibition by quinone binding site tool compounds. The setup relies on their immobilization at electrodes specifically modified with gold nanoparticles, which allows achieving a direct electron transfer to/from the heme cofactors of this large enzyme. After optimization of the protein coverages, the assay shows at pH 7 a good reproducibility and readout stability over time, and it is thus suitable for further screening of small molecule collections.

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