The YheI/YheH heterodimer from Bacillus subtilis is a multidrug ABC transporter |
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Authors: | Cristina Torres Carmen Galián Christoph Freiberg Jean-Michel Jault |
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Institution: | a Institut de Biologie Structurale, UMR 5075 Université Joseph Fourier/CEA/CNRS, 41 rue Jules Horowitz 38027 Grenoble cedex 1, France b Niederrhein University of Applied Sciences, Department of Chemistry, Adlerstrasse 32, D-47798 Krefeld, Germany c Laboratoire de Chimie Bactérienne, UPR 9043 du CNRS, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France |
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Abstract: | ABC (ATP-binding cassette) transporters form the largest family of membrane proteins in micro-organisms where they are able to transport a wide variety of substrates against a concentration gradient, in an ATP-dependent process. Two genes from the same putative Bacillus subtilis operon, yheI and yheH, encoding possibly two different ABC transporters, were overexpressed in Escherichia coli in high yield, either separately or jointly. Using membrane vesicles, it is shown here that both subunits were required to detect, (i) the transport of four structurally unrelated drugs, and (ii) a vanadate-sensitive ATPase activity. Mutation of the invariant Walker-A lysine to an alanine residue in both subunits led to an inactive transporter. Moreover, after membrane solubilization by detergent, both wild-type subunits co-purified on a Ni-Agarose affinity column while only the YheH subunit contained a hexa-histidine tag. This shows that YheI and YheH are indeed able to interact together to form a heterodimer. Importantly, expression of both yheI and yheH genes in B. subtilis could be strongly stimulated by addition of sub-inhibitory concentrations of various unrelated antibiotics. Therefore, B. subtilis YheI/YheH forms a new heterodimeric multidrug ABC transporter possibly involved in multiple antibiotic resistance in vivo. |
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Keywords: | ABC transporter Multidrug Overexpression Drug transport ATP-binding cassette |
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