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Fitting periplasmic membrane fusion proteins to inner membrane transporters: mutations that enable Escherichia coli AcrA to function with Pseudomonas aeruginosa MexB.
J Bacteriol. 2008 Jan; 190(2):691-8.JB

Abstract

AcrAB-TolC from Escherichia coli is a multidrug efflux complex capable of transenvelope transport. In this complex, AcrA is a periplasmic membrane fusion protein that establishes a functional connection between the inner membrane transporter AcrB of the RND superfamily and the outer membrane channel TolC. To gain insight into the mechanism of the functional association between components of this complex, we replaced AcrB with its close homolog MexB from Pseudomonas aeruginosa. Surprisingly, we found that AcrA is promiscuous and can form a partially functional complex with MexB and TolC. The chimeric AcrA-MexB-TolC complex protected cells from sodium dodecyl sulfate, novobiocin, and ethidium bromide but failed with other known substrates of MexB. We next identified single and double mutations in AcrA and MexB that enabled the complete functional fit between AcrA, MexB, and TolC. Mutations in either the alpha-helical hairpin of AcrA making contact with TolC or the beta-barrel domain lying on MexB improved the functional alignment between components of the complex. Our results suggest that three components of multidrug efflux pumps do not associate in an "all-or-nothing" fashion but accommodate a certain degree of flexibility. This flexibility in the association between components affects the transport efficiency of RND pumps.

Authors+Show Affiliations

Department of Chemistry and Biochemistry, University of Oklahoma, 620 Parrington Oval, Room 208, Norman, OK 73019, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural

Language

eng

PubMed ID

18024521

Citation

Krishnamoorthy, Ganesh, et al. "Fitting Periplasmic Membrane Fusion Proteins to Inner Membrane Transporters: Mutations That Enable Escherichia Coli AcrA to Function With Pseudomonas Aeruginosa MexB." Journal of Bacteriology, vol. 190, no. 2, 2008, pp. 691-8.
Krishnamoorthy G, Tikhonova EB, Zgurskaya HI. Fitting periplasmic membrane fusion proteins to inner membrane transporters: mutations that enable Escherichia coli AcrA to function with Pseudomonas aeruginosa MexB. J Bacteriol. 2008;190(2):691-8.
Krishnamoorthy, G., Tikhonova, E. B., & Zgurskaya, H. I. (2008). Fitting periplasmic membrane fusion proteins to inner membrane transporters: mutations that enable Escherichia coli AcrA to function with Pseudomonas aeruginosa MexB. Journal of Bacteriology, 190(2), 691-8.
Krishnamoorthy G, Tikhonova EB, Zgurskaya HI. Fitting Periplasmic Membrane Fusion Proteins to Inner Membrane Transporters: Mutations That Enable Escherichia Coli AcrA to Function With Pseudomonas Aeruginosa MexB. J Bacteriol. 2008;190(2):691-8. PubMed PMID: 18024521.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Fitting periplasmic membrane fusion proteins to inner membrane transporters: mutations that enable Escherichia coli AcrA to function with Pseudomonas aeruginosa MexB. AU - Krishnamoorthy,Ganesh, AU - Tikhonova,Elena B, AU - Zgurskaya,Helen I, Y1 - 2007/11/16/ PY - 2007/11/21/pubmed PY - 2008/2/2/medline PY - 2007/11/21/entrez SP - 691 EP - 8 JF - Journal of bacteriology JO - J. Bacteriol. VL - 190 IS - 2 N2 - AcrAB-TolC from Escherichia coli is a multidrug efflux complex capable of transenvelope transport. In this complex, AcrA is a periplasmic membrane fusion protein that establishes a functional connection between the inner membrane transporter AcrB of the RND superfamily and the outer membrane channel TolC. To gain insight into the mechanism of the functional association between components of this complex, we replaced AcrB with its close homolog MexB from Pseudomonas aeruginosa. Surprisingly, we found that AcrA is promiscuous and can form a partially functional complex with MexB and TolC. The chimeric AcrA-MexB-TolC complex protected cells from sodium dodecyl sulfate, novobiocin, and ethidium bromide but failed with other known substrates of MexB. We next identified single and double mutations in AcrA and MexB that enabled the complete functional fit between AcrA, MexB, and TolC. Mutations in either the alpha-helical hairpin of AcrA making contact with TolC or the beta-barrel domain lying on MexB improved the functional alignment between components of the complex. Our results suggest that three components of multidrug efflux pumps do not associate in an "all-or-nothing" fashion but accommodate a certain degree of flexibility. This flexibility in the association between components affects the transport efficiency of RND pumps. SN - 1098-5530 UR - https://www.unboundmedicine.com/medline/citation/18024521/Fitting_periplasmic_membrane_fusion_proteins_to_inner_membrane_transporters:_mutations_that_enable_Escherichia_coli_AcrA_to_function_with_Pseudomonas_aeruginosa_MexB_ L2 - http://jb.asm.org/cgi/pmidlookup?view=long&pmid=18024521 DB - PRIME DP - Unbound Medicine ER -