Activity Monitoring of Functional OprM Using a Biomimetic Microfluidic Device Article - Janvier 2012

Wei Wang, Laura Monlezun, Martin Picard, Philippe Benas, Olivier Français, Isabelle Broutin, Bruno Le Pioufle

Wei Wang, Laura Monlezun, Martin Picard, Philippe Benas, Olivier Français, Isabelle Broutin, Bruno Le Pioufle, « Activity Monitoring of Functional OprM Using a Biomimetic Microfluidic Device  », Analyst, janvier 2012, pp. 847-852. ISSN 0003-2654

Abstract

This paper describes the fabrication and use of a biomimetic microfluidic device for the monitoring of a functional porin reconstituted within miniaturized suspended artificial bilayer lipid membrane (BLM). Such a microfluidic device allows for 1) fluidic and electrical access to both sides of the BLM, 2) reproducible membrane protein insertion and long-term electrical monitoring of its conductance (Gi), thanks to the miniaturization of the BLM. We demonstrate here for the first time the feasibility to insert a large trans-membrane protein through its β-barrel, and monitor its functional activity during more than 1 hour (limited by buffer evaporation). In this paper, we specifically used our device for the monitoring of OprM, a bacterial efflux channel involved in the multidrug resistance of the bacteria Pseudomonas aeruginosa. Sub-steps of the OprM channel conductance were detected during the electrical recordings within our device, which might be due to oscillations between several structural conformations (sub-states) adopted by the protein, as part of its opening mechanism. This work is a first step towards the establishing of a genuine platform dedicated to the investigation of bacterial proteins under reconstituted conditions, a very promising tool for the screening of new inhibitors against bacterial channels involved in drug resistance.

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