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- Publisher Website: 10.1016/j.jmb.2011.05.042
- Scopus: eid_2-s2.0-79960699901
- PMID: 21664361
- WOS: WOS:000293938300019
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Article: Folding of AcrB subunit precedes trimerization
Title | Folding of AcrB subunit precedes trimerization |
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Authors | |
Keywords | folding and assembly membrane protein oligomerization obligate oligomer protein tertiary and quaternary structures thiol trapping |
Issue Date | 2011 |
Publisher | Academic 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? |
Abstract | AcrB 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 Identifier | http://hdl.handle.net/10722/168545 |
ISSN | 2023 Impact Factor: 4.7 2023 SCImago Journal Rankings: 2.212 |
ISI Accession Number ID | |
References |
DC Field | Value | Language |
---|---|---|
dc.contributor.author | Lu, W | en_US |
dc.contributor.author | Zhong, M | en_US |
dc.contributor.author | Wei, Y | en_US |
dc.date.accessioned | 2012-10-08T03:20:23Z | - |
dc.date.available | 2012-10-08T03:20:23Z | - |
dc.date.issued | 2011 | en_US |
dc.identifier.citation | Journal Of Molecular Biology, 2011, v. 411 n. 1, p. 264-274 | en_US |
dc.identifier.issn | 0022-2836 | en_US |
dc.identifier.uri | http://hdl.handle.net/10722/168545 | - |
dc.description.abstract | AcrB 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.language | eng | en_US |
dc.publisher | Academic Press. The Journal's web site is located at http://www.elsevier.com/locate/jmb | en_US |
dc.relation.ispartof | Journal of Molecular Biology | en_US |
dc.subject | folding and assembly | - |
dc.subject | membrane protein oligomerization | - |
dc.subject | obligate oligomer | - |
dc.subject | protein tertiary and quaternary structures | - |
dc.subject | thiol trapping | - |
dc.subject.mesh | Anti-Bacterial Agents - Pharmacology | en_US |
dc.subject.mesh | Electrophoresis, Polyacrylamide Gel | en_US |
dc.subject.mesh | Escherichia Coli - Drug Effects - Genetics - Metabolism | en_US |
dc.subject.mesh | Escherichia Coli Proteins - Chemistry - Genetics - Metabolism | en_US |
dc.subject.mesh | Microbial Sensitivity Tests | en_US |
dc.subject.mesh | Models, Molecular | en_US |
dc.subject.mesh | Multidrug Resistance-Associated Proteins - Chemistry - Genetics - Metabolism | en_US |
dc.subject.mesh | Mutant Proteins - Genetics - Metabolism | en_US |
dc.subject.mesh | Protein Folding | en_US |
dc.subject.mesh | Protein Multimerization | en_US |
dc.subject.mesh | Protein Structure, Quaternary | en_US |
dc.subject.mesh | Protein Structure, Tertiary | en_US |
dc.subject.mesh | Protein Subunits - Chemistry - Genetics - Metabolism | en_US |
dc.subject.mesh | Sequence Deletion | en_US |
dc.title | Folding of AcrB subunit precedes trimerization | en_US |
dc.type | Article | en_US |
dc.identifier.email | Lu, W:luwei@hku.hk | en_US |
dc.identifier.authority | Lu, W=rp00754 | en_US |
dc.description.nature | link_to_subscribed_fulltext | en_US |
dc.identifier.doi | 10.1016/j.jmb.2011.05.042 | en_US |
dc.identifier.pmid | 21664361 | en_US |
dc.identifier.scopus | eid_2-s2.0-79960699901 | en_US |
dc.relation.references | http://www.scopus.com/mlt/select.url?eid=2-s2.0-79960699901&selection=ref&src=s&origin=recordpage | en_US |
dc.identifier.volume | 411 | en_US |
dc.identifier.issue | 1 | en_US |
dc.identifier.spage | 264 | en_US |
dc.identifier.epage | 274 | en_US |
dc.identifier.isi | WOS:000293938300019 | - |
dc.publisher.place | United Kingdom | en_US |
dc.identifier.scopusauthorid | Lu, W=27868087600 | en_US |
dc.identifier.scopusauthorid | Zhong, M=36184255700 | en_US |
dc.identifier.scopusauthorid | Wei, Y=7404094290 | en_US |
dc.identifier.citeulike | 9414601 | - |
dc.identifier.issnl | 0022-2836 | - |