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E. coli Multidrug Transporter MdfA Is a Monomer

Sigal, Nadejda and Lewinson, Oded and Wolf, Sharon G. and Bibi, Eitan (2007) E. coli Multidrug Transporter MdfA Is a Monomer. Biochemistry, 46 (17). pp. 5200-5208. ISSN 0006-2960. doi:10.1021/bi602405w. https://resolver.caltech.edu/CaltechAUTHORS:20170131-140942712

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Abstract

MdfA is a 410-residue-long secondary multidrug transporter from E. coli. Cells expressing MdfA from a multicopy plasmid exhibit resistance against a diverse group of toxic compounds, including neutral and cationic ones, because of active multidrug export. As a prerequisite for high-resolution structural studies and a better understanding of the mechanism of substrate recognition and translocation by MdfA, we investigated its biochemical properties and overall structural characteristics. To this end, we purified the β-dodecyl maltopyranoside (DDM)-solubilized protein using a 6-His tag and metal affinity chromatography, and size exclusion chromatography (SE-HPLC). Purified MdfA was analyzed for its DDM and phospholipid (PL) content, and tetraphenylphosphonium (TPP+)-binding activity. The results are consistent with MdfA being an active monomer in DDM solution. Furthermore, an investigation of two-dimensional crystals by electron crystallography and 3D reconstruction lent support to the notion that MdfA may also be monomeric in reconstituted proteoliposomes.


Item Type:Article
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URLURL TypeDescription
http://dx.doi.org/10.1021/bi602405wDOIArticle
http://pubs.acs.org/doi/abs/10.1021/bi602405wPublisherArticle
Additional Information:© 2007 American Chemical Society. Received 21 November 2006. Published online 4 April 2007. Published in print 1 May 2007. This work was supported by a grant from the Israel Cancer Research Foundation (ICRF) and the Israel Science Foundation (ISF). We thank Werner Kuhlbrandt and Thomas Walz for their help and advice in the initial stages of the 2D crystallization project, Daniel Tal for his help during the SE-HPLC studies, and Lubov Ginzburg for help in the phospholipid assays.
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Funding AgencyGrant Number
Israel Cancer Research FoundationUNSPECIFIED
Israel Science FoundationUNSPECIFIED
Issue or Number:17
DOI:10.1021/bi602405w
Record Number:CaltechAUTHORS:20170131-140942712
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20170131-140942712
Official Citation:E. coli Multidrug Transporter MdfA Is a Monomer Nadejda Sigal, Oded Lewinson, Sharon G. Wolf, and Eitan Bibi Biochemistry 2007 46 (17), 5200-5208 DOI: 10.1021/bi602405w
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:73896
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:01 Feb 2017 00:33
Last Modified:11 Nov 2021 05:23

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