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Article: Folding of AcrB subunit precedes trimerization

TitleFolding of AcrB subunit precedes trimerization
Authors
Issue Date2011
PublisherAcademic Press. The Journal's web site is located at http://www.elsevier.com/locate/jmb
Citation
Journal Of Molecular Biology, 2011, v. 411 n. 1, p. 264-274 How to Cite?
AbstractAcrB and its homologues are major players in the efflux of anti-microbials out of Gram-negative bacteria. The structural and functional unit of AcrB is a homo-trimer. The assembly process of obligate membrane protein oligomers, including AcrB, remains elusive. It is not clear if an individual subunit folds into a monomeric form first followed by association (three-stage pathway) or if association occurs simultaneously with subunit folding (two-stage pathway). To answer this question, we investigated the feasibility of creating a folded monomeric AcrB mutant. The existence of well-folded monomers in the cell membrane would be an evidence of a three-stage pathway. A monomeric AcrB mutant, AcrB Δloop, was created through the truncation of a protruding loop that appeared to contribute to the stability of an AcrB trimer. AcrB Δloop expressed at a level similar to that of wild-type AcrB. The secondary structure content and tertiary conformation of AcrB Δloop were very similar to those of wild-type AcrB. However, when expressed in an acrB-deficient strain, AcrB Δloop failed to complement its defect in drug efflux. Results from blue native polyacrylamide gel electrophoresis and chemical cross-linking experiments suggested that AcrB Δloop existed as a monomer. The expression of this monomeric mutant in a wild-type Escherichia coli strain did not have a significant dominant-negative effect, suggesting that the mutant could not effectively co-assemble with genomic AcrB. AcrB Δloop is the first monomeric mutant reported for the intrinsically trimeric AcrB. The structural characterization results of this mutant suggest that the oligomerization of AcrB occurs through a three-stage pathway involving folded monomers. © 2011 Elsevier Ltd. All rights reserved.
Persistent Identifierhttp://hdl.handle.net/10722/168545
ISSN
2015 Impact Factor: 4.517
2015 SCImago Journal Rankings: 3.002
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorLu, Wen_US
dc.contributor.authorZhong, Men_US
dc.contributor.authorWei, Yen_US
dc.date.accessioned2012-10-08T03:20:23Z-
dc.date.available2012-10-08T03:20:23Z-
dc.date.issued2011en_US
dc.identifier.citationJournal Of Molecular Biology, 2011, v. 411 n. 1, p. 264-274en_US
dc.identifier.issn0022-2836en_US
dc.identifier.urihttp://hdl.handle.net/10722/168545-
dc.description.abstractAcrB and its homologues are major players in the efflux of anti-microbials out of Gram-negative bacteria. The structural and functional unit of AcrB is a homo-trimer. The assembly process of obligate membrane protein oligomers, including AcrB, remains elusive. It is not clear if an individual subunit folds into a monomeric form first followed by association (three-stage pathway) or if association occurs simultaneously with subunit folding (two-stage pathway). To answer this question, we investigated the feasibility of creating a folded monomeric AcrB mutant. The existence of well-folded monomers in the cell membrane would be an evidence of a three-stage pathway. A monomeric AcrB mutant, AcrB Δloop, was created through the truncation of a protruding loop that appeared to contribute to the stability of an AcrB trimer. AcrB Δloop expressed at a level similar to that of wild-type AcrB. The secondary structure content and tertiary conformation of AcrB Δloop were very similar to those of wild-type AcrB. However, when expressed in an acrB-deficient strain, AcrB Δloop failed to complement its defect in drug efflux. Results from blue native polyacrylamide gel electrophoresis and chemical cross-linking experiments suggested that AcrB Δloop existed as a monomer. The expression of this monomeric mutant in a wild-type Escherichia coli strain did not have a significant dominant-negative effect, suggesting that the mutant could not effectively co-assemble with genomic AcrB. AcrB Δloop is the first monomeric mutant reported for the intrinsically trimeric AcrB. The structural characterization results of this mutant suggest that the oligomerization of AcrB occurs through a three-stage pathway involving folded monomers. © 2011 Elsevier Ltd. All rights reserved.en_US
dc.languageengen_US
dc.publisherAcademic Press. The Journal's web site is located at http://www.elsevier.com/locate/jmben_US
dc.relation.ispartofJournal of Molecular Biologyen_US
dc.subject.meshAnti-Bacterial Agents - Pharmacologyen_US
dc.subject.meshElectrophoresis, Polyacrylamide Gelen_US
dc.subject.meshEscherichia Coli - Drug Effects - Genetics - Metabolismen_US
dc.subject.meshEscherichia Coli Proteins - Chemistry - Genetics - Metabolismen_US
dc.subject.meshMicrobial Sensitivity Testsen_US
dc.subject.meshModels, Molecularen_US
dc.subject.meshMultidrug Resistance-Associated Proteins - Chemistry - Genetics - Metabolismen_US
dc.subject.meshMutant Proteins - Genetics - Metabolismen_US
dc.subject.meshProtein Foldingen_US
dc.subject.meshProtein Multimerizationen_US
dc.subject.meshProtein Structure, Quaternaryen_US
dc.subject.meshProtein Structure, Tertiaryen_US
dc.subject.meshProtein Subunits - Chemistry - Genetics - Metabolismen_US
dc.subject.meshSequence Deletionen_US
dc.titleFolding of AcrB subunit precedes trimerizationen_US
dc.typeArticleen_US
dc.identifier.emailLu, W:luwei@hku.hken_US
dc.identifier.authorityLu, W=rp00754en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1016/j.jmb.2011.05.042en_US
dc.identifier.pmid21664361en_US
dc.identifier.scopuseid_2-s2.0-79960699901en_US
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-79960699901&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume411en_US
dc.identifier.issue1en_US
dc.identifier.spage264en_US
dc.identifier.epage274en_US
dc.identifier.isiWOS:000293938300019-
dc.publisher.placeUnited Kingdomen_US
dc.identifier.scopusauthoridLu, W=27868087600en_US
dc.identifier.scopusauthoridZhong, M=36184255700en_US
dc.identifier.scopusauthoridWei, Y=7404094290en_US
dc.identifier.citeulike9414601-

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