Opening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partner

2.50
Hdl Handle:
http://hdl.handle.net/10547/622750
Title:
Opening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partner
Authors:
Janganan, Thamarai K.; Zhang, Li; Bavro, Vassiliy N ( 0000-0003-2330-8924 ) ; Matak-Vinkovic, Dijana; Barrera, Nelson P.; Burton, Matthew F.; Steel, Patrick G.; Robinson, Carol V.; Borges-Walmsley, Maria Ines; Walmsley, Adrian R. ( 0000-0002-5136-0511 )
Abstract:
The multiple transferable resistance (MTR) pump, from Neisseria gonorrhoeae, is typical of the specialized machinery used to translocate drugs across the inner and outer membranes of Gram-negative bacteria. It consists of a tripartite complex composed of an inner-membrane transporter, MtrD, a periplasmic membrane fusion protein, MtrC, and an outer-membrane channel, MtrE. We have expressed the components of the pump in Escherichia coli and used the antibiotic vancomycin, which is too large to cross the outer-membrane by passive diffusion, to test for opening of the MtrE channel. Cells expressing MtrCDE are not susceptible to vancomycin, indicating that the channel is closed; but become susceptible to vancomycin in the presence of transported substrates, consistent with drug-induced opening of the MtrE channel. A mutational analysis identified residues Asn-198, Glu-434, and Gln-441, lining an intraprotomer groove on the surface of MtrE, to be important for pump function; mutation of these residues yielded cells that were sensitive to vancomycin. Pull-down assays and micro-calorimetry measurements indicated that this functional impairment is not due to the inability of MtrC to interact with the MtrE mutants; nor was it due to the MtrE mutants adopting an open conformation, because cells expressing these MtrE mutants alone are relatively insensitive to vancomycin. However, cells expressing the MtrE mutants with MtrC are sensitive to vancomycin, indicating that residues lining the intra-protomer groove control opening of the MtrE channel in response to binding of MtrC.
Affiliation:
University of Durham; University of Oxford; University of Cambridge
Citation:
Janganan TL, Zhang Li, Bavro VN, Matak-Vinkovic D, Barrera NP, Burton MF, Steel PG, Robinson CV, Borges-Walmsley, Walmsley AR (2011) 'Opening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partner.', Journal of Biological Chemistry, 286 (7), pp.5484-93.
Publisher:
American Society for Biochemistry and Molecular Biology
Journal:
Journal of Biological Chemistry
Issue Date:
18-Feb-2011
URI:
http://hdl.handle.net/10547/622750
DOI:
10.1074/jbc.M110.187658
PubMed ID:
21115481
Additional Links:
http://www.jbc.org/content/286/7/5484.short
Type:
Article
Language:
en
ISSN:
0021-9258
Appears in Collections:
Biomedical and biological science

Full metadata record

DC FieldValue Language
dc.contributor.authorJanganan, Thamarai K.en
dc.contributor.authorZhang, Lien
dc.contributor.authorBavro, Vassiliy Nen
dc.contributor.authorMatak-Vinkovic, Dijanaen
dc.contributor.authorBarrera, Nelson P.en
dc.contributor.authorBurton, Matthew F.en
dc.contributor.authorSteel, Patrick G.en
dc.contributor.authorRobinson, Carol V.en
dc.contributor.authorBorges-Walmsley, Maria Inesen
dc.contributor.authorWalmsley, Adrian R.en
dc.date.accessioned2018-06-20T11:00:14Z-
dc.date.available2018-06-20T11:00:14Z-
dc.date.issued2011-02-18-
dc.identifier.citationJanganan TL, Zhang Li, Bavro VN, Matak-Vinkovic D, Barrera NP, Burton MF, Steel PG, Robinson CV, Borges-Walmsley, Walmsley AR (2011) 'Opening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partner.', Journal of Biological Chemistry, 286 (7), pp.5484-93.en
dc.identifier.issn0021-9258-
dc.identifier.pmid21115481-
dc.identifier.doi10.1074/jbc.M110.187658-
dc.identifier.urihttp://hdl.handle.net/10547/622750-
dc.description.abstractThe multiple transferable resistance (MTR) pump, from Neisseria gonorrhoeae, is typical of the specialized machinery used to translocate drugs across the inner and outer membranes of Gram-negative bacteria. It consists of a tripartite complex composed of an inner-membrane transporter, MtrD, a periplasmic membrane fusion protein, MtrC, and an outer-membrane channel, MtrE. We have expressed the components of the pump in Escherichia coli and used the antibiotic vancomycin, which is too large to cross the outer-membrane by passive diffusion, to test for opening of the MtrE channel. Cells expressing MtrCDE are not susceptible to vancomycin, indicating that the channel is closed; but become susceptible to vancomycin in the presence of transported substrates, consistent with drug-induced opening of the MtrE channel. A mutational analysis identified residues Asn-198, Glu-434, and Gln-441, lining an intraprotomer groove on the surface of MtrE, to be important for pump function; mutation of these residues yielded cells that were sensitive to vancomycin. Pull-down assays and micro-calorimetry measurements indicated that this functional impairment is not due to the inability of MtrC to interact with the MtrE mutants; nor was it due to the MtrE mutants adopting an open conformation, because cells expressing these MtrE mutants alone are relatively insensitive to vancomycin. However, cells expressing the MtrE mutants with MtrC are sensitive to vancomycin, indicating that residues lining the intra-protomer groove control opening of the MtrE channel in response to binding of MtrC.en
dc.language.isoenen
dc.publisherAmerican Society for Biochemistry and Molecular Biologyen
dc.relation.urlhttp://www.jbc.org/content/286/7/5484.shorten
dc.rightsGreen - can archive pre-print and post-print or publisher's version/PDF-
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/*
dc.subjecttripartite efflux pumpen
dc.subjectmultidrug transportersen
dc.subjectNeisseria gonorrhoeaeen
dc.titleOpening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partneren
dc.typeArticleen
dc.contributor.departmentUniversity of Durhamen
dc.contributor.departmentUniversity of Oxforden
dc.contributor.departmentUniversity of Cambridgeen
dc.identifier.journalJournal of Biological Chemistryen
dc.date.updated2018-06-20T09:02:43Z-

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