Overexpression of the ABC Transporter BmrA Within Intracellular Caveolae in Escherichia coli

• DOI: 10.1007/978-1-0716-2368-8_5
• Publication Date: 2022-06-30T16:21:33Z
• Authors: Alexia Gobet; Christine Jaxel; Sandrine Magnard; Manuel Garrigos; Stéphane Orlowski; Nadège Jamin; Pierre Falson; Vincent Chaptal
• Keywords: Efflux; Caveolin; Heterologous expression

We describe here the overproduction and oriented membrane insertion of membrane protein inside intracellular vesicles named heterologous caveolae within E. coli. The method is described with BmrA, a multidrug efflux pump from Bacillus subtilis. BmrA is produced in these vesicles, thanks to the coexpression with the canine caveolin-1β, one of the two isoforms of caveolin-1. Enriched by sucrose gradient, the caveolae-containing fraction allows to probe the ATPase and Hoechst 33342 transport activities, the latter displaying a higher specific activity than the same without caveolin-1β.