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PDBsum entry 2v4d

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protein ligands Protein-protein interface(s) links
Membrane protein PDB id
2v4d
Jmol
Contents
Protein chains
232 a.a. *
(+ 5 more) 327 a.a. *
Ligands
SO4 ×16
* Residue conservation analysis
PDB id:
2v4d
Name: Membrane protein
Title: Re-refinement of mexa adaptor protein
Structure: Multidrug resistance protein mexa. Chain: a, b, c, d, e, f, g, h, i, j, k, l, m. Engineered: yes. Mutation: yes
Source: Pseudomonas aeruginosa. Organism_taxid: 287. Expressed in: escherichia coli. Expression_system_taxid: 562
Resolution:
3.20Å     R-factor:   0.240     R-free:   0.264
Authors: M.F.Symmons,E.Bokma,E.Koronakis,C.Hughes,V.Koronakis
Key ref:
M.F.Symmons et al. (2009). The assembled structure of a complete tripartite bacterial multidrug efflux pump. Proc Natl Acad Sci U S A, 106, 7173-7178. PubMed id: 19342493 DOI: 10.1073/pnas.0900693106
Date:
18-Sep-08     Release date:   14-Apr-09    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P52477  (MEXA_PSEAE) -  Multidrug resistance protein MexA
Seq:
Struc:
383 a.a.
232 a.a.
Protein chains
Pfam   ArchSchema ?
P52477  (MEXA_PSEAE) -  Multidrug resistance protein MexA
Seq:
Struc:
383 a.a.
327 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   2 terms 
  Biological process     transport   4 terms 
  Biochemical function     drug transmembrane transporter activity     1 term  

 

 
DOI no: 10.1073/pnas.0900693106 Proc Natl Acad Sci U S A 106:7173-7178 (2009)
PubMed id: 19342493  
 
 
The assembled structure of a complete tripartite bacterial multidrug efflux pump.
M.F.Symmons, E.Bokma, E.Koronakis, C.Hughes, V.Koronakis.
 
  ABSTRACT  
 
Bacteria like Escherichia coli and Pseudomonas aeruginosa expel drugs via tripartite multidrug efflux pumps spanning both inner and outer membranes and the intervening periplasm. In these pumps a periplasmic adaptor protein connects a substrate-binding inner membrane transporter to an outer membrane-anchored TolC-type exit duct. High-resolution structures of all 3 components are available, but a pump model has been precluded by the incomplete adaptor structure, because of the apparent disorder of its N and C termini. We reveal that the adaptor termini assemble a beta-roll structure forming the final domain adjacent to the inner membrane. The completed structure enabled in vivo cross-linking to map intermolecular contacts between the adaptor AcrA and the transporter AcrB, defining a periplasmic interface between several transporter subdomains and the contiguous beta-roll, beta-barrel, and lipoyl domains of the adaptor. With short and long cross-links expressed as distance restraints, the flexible linear topology of the adaptor allowed a multidomain docking approach to model the transporter-adaptor complex, revealing that the adaptor docks to a transporter region of comparative stability distinct from those key to the proposed rotatory pump mechanism, putative drug-binding pockets, and the binding site of inhibitory DARPins. Finally, we combined this docking with our previous resolution of the AcrA hairpin-TolC interaction to develop a model of the assembled tripartite complex, satisfying all of the experimentally-derived distance constraints. This AcrA(3)-AcrB(3)-TolC(3) model presents a 610,000-Da, 270-A-long efflux pump crossing the entire bacterial cell envelope.
 
  Selected figure(s)  
 
Figure 2.
The completed structure of the periplasmic adaptor. (A) Structure of MexA including the MP domain. The 4 adaptor domains are: α-hairpin (blue), lipoyl (green), β-barrel (yellow), and MP β-roll (orange). Turns are gray except for 2 MP domain helical turns (yellow) that include Gly residues (white Cα atoms) on the concave surface effecting crystal contacts. The enlarged inset gives a smoothed representation of the MP domain topology, with elements numbered according to the adaptor family sequence alignment (Fig. S1) and colored from blue to red. Trp-309 is shown in gray. (B) The MP domain: conformational variation, rotation, and crystal contacts. The MexA adaptor barrel (yellow) and MP (orange) domain, shown in the unrotated MP domain conformation, establish crystal contacts with a neighboring copy (gray), with the MP domain in its rotated conformation. Helical turns on the MP domain concave face are in yellow, and Gly-281 and Trp-309 are shown as white Cα atoms and gray side-chains, respectively (labeled in italics on the rotated domain).
Figure 4.
Docking of AcrA and AcrB directed by the cross-linking data. (A) AcrA and AcrB interaction hotspots predocking. Cross-linking hotspots on the AcrA and AcrB surfaces predocking, colored on a gradient reflecting all cross-linking data: from dark blue near negative Cys-substituted residues (no cross-link), through green near residues with a bias to the L linker, to yellow/orange/red with increasing proximity to positives linked by both S and L linker. The TolC interface of the adaptor hairpin (23) is in magenta. The arrow indicates rotation from side to front view. (B) Docked complex of AcrA on an AcrB subunit. The 4 AcrA adaptor domains are shown in green shades, with 2 helical turns of the MP concave surface in yellow, β-turn Gly residues in white and the N-terminal residue in blue. The transporter periplasmic subdomain colors are as in Fig. 3B. Cross-sections at 2 levels of the complex (indicated by the brackets) are boxed on the right and illustrate the domain–domain contacts of the adaptor MP domain (i) and β-barrel and lipoyl domains (ii).
 
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20981744 C.C.Su, F.Long, and E.W.Yu (2011).
The Cus efflux system removes toxic ions via a methionine shuttle.
  Protein Sci, 20, 6.  
21350490 C.C.Su, F.Long, M.T.Zimmermann, K.R.Rajashankar, R.L.Jernigan, and E.W.Yu (2011).
Crystal structure of the CusBA heavy-metal efflux complex of Escherichia coli.
  Nature, 470, 558-562.
PDB code: 3ne5
21513873 C.Oswald, and K.M.Pos (2011).
Drug resistance: a periplasmic ménage à trois.
  Chem Biol, 18, 405-407.  
21513882 E.B.Tikhonova, Y.Yamada, and H.I.Zgurskaya (2011).
Sequential mechanism of assembly of multidrug efflux pump AcrAB-TolC.
  Chem Biol, 18, 454-463.  
21549704 J.M.Bolla, S.Alibert-Franco, J.Handzlik, J.Chevalier, A.Mahamoud, G.Boyer, K.Kieć-Kononowicz, and J.M.Pagès (2011).
Strategies for bypassing the membrane barrier in multidrug resistant Gram-negative bacteria.
  FEBS Lett, 585, 1682-1690.  
21296164 N.Monroe, G.Sennhauser, M.A.Seeger, C.Briand, and M.G.Grütter (2011).
Designed ankyrin repeat protein binders for the crystallization of AcrB: plasticity of the dominant interface.
  J Struct Biol, 174, 269-281.
PDB codes: 3noc 3nog
21249122 R.Kulathila, R.Kulathila, M.Indic, and B.van den Berg (2011).
Crystal structure of Escherichia coli CusC, the outer membrane component of a heavy metal efflux pump.
  PLoS One, 6, e15610.
PDB code: 3pik
22121023 R.Nakashima, K.Sakurai, S.Yamasaki, K.Nishino, and A.Yamaguchi (2011).
Structures of the multidrug exporter AcrB reveal a proximal multisite drug-binding pocket.
  Nature, 480, 565-569.
PDB codes: 3aoa 3aob 3aoc 3aod
21115481 T.K.Janganan, L.Zhang, V.N.Bavro, D.Matak-Vinkovic, N.P.Barrera, M.F.Burton, P.G.Steel, C.V.Robinson, M.I.Borges-Walmsley, and A.R.Walmsley (2011).
Opening of the outer membrane protein channel in tripartite efflux pumps is induced by interaction with the membrane fusion partner.
  J Biol Chem, 286, 5484-5493.  
21371926 Y.S.Choong, T.S.Lim, A.L.Chew, I.Aziah, and A.Ismail (2011).
Structural and functional studies of a 50 kDa antigenic protein from Salmonella enterica serovar Typhi.
  J Mol Graph Model, 29, 834-842.  
20442961 E.H.Kim, C.Rensing, and M.M.McEvoy (2010).
Chaperone-mediated copper handling in the periplasm.
  Nat Prod Rep, 27, 711-719.  
20534468 F.De Angelis, J.K.Lee, J.D.O'Connell, L.J.Miercke, K.H.Verschueren, V.Srinivasan, C.Bauvois, C.Govaerts, R.A.Robbins, J.M.Ruysschaert, R.M.Stroud, and G.Vandenbussche (2010).
Metal-induced conformational changes in ZneB suggest an active role of membrane fusion proteins in efflux resistance systems.
  Proc Natl Acad Sci U S A, 107, 11038-11043.
PDB code: 3lnn
20865003 F.Long, C.C.Su, M.T.Zimmermann, S.E.Boyken, K.R.Rajashankar, R.L.Jernigan, and E.W.Yu (2010).
Crystal structures of the CusA efflux pump suggest methionine-mediated metal transport.
  Nature, 467, 484-488.
PDB codes: 3k07 3kso 3kss
20581201 H.M.Kim, Y.Xu, M.Lee, S.Piao, S.H.Sim, N.C.Ha, and K.Lee (2010).
Functional relationships between the AcrA hairpin tip region and the TolC aperture tip region for the formation of the bacterial tripartite efflux pump AcrAB-TolC.
  J Bacteriol, 192, 4498-4503.  
20038594 H.S.Kim, D.Nagore, and H.Nikaido (2010).
Multidrug efflux pump MdtBC of Escherichia coli is active only as a B2C heterotrimer.
  J Bacteriol, 192, 1377-1386.  
20606071 J.A.Bohnert, B.Karamian, and H.Nikaido (2010).
Optimized Nile Red efflux assay of AcrAB-TolC multidrug efflux system shows competition between substrates.
  Antimicrob Agents Chemother, 54, 3770-3775.  
20132445 J.W.Weeks, T.Celaya-Kolb, S.Pecora, and R.Misra (2010).
AcrA suppressor alterations reverse the drug hypersensitivity phenotype of a TolC mutant by inducing TolC aperture opening.
  Mol Microbiol, 75, 1468-1483.  
20805930 M.J.Dunlop, J.D.Keasling, and A.Mukhopadhyay (2010).
A model for improving microbial biofuel production using a synthetic feedback loop.
  Syst Synth Biol, 4, 95.  
20548943 R.Schulz, A.V.Vargiu, F.Collu, U.Kleinekathöfer, and P.Ruggerone (2010).
Functional rotation of the transporter AcrB: insights into drug extrusion from simulations.
  PLoS Comput Biol, 6, e1000806.  
20160052 S.P.Lim, and H.Nikaido (2010).
Kinetic parameters of efflux of penicillins by the multidrug efflux transporter AcrAB-TolC of Escherichia coli.
  Antimicrob Agents Chemother, 54, 1800-1806.  
  20452953 T.J.Silhavy, D.Kahne, and S.Walker (2010).
The bacterial cell envelope.
  Cold Spring Harb Perspect Biol, 2, a000414.  
  21081915 X.Q.Yao, H.Kenzaki, S.Murakami, and S.Takada (2010).
Drug export and allosteric coupling in a multidrug transporter revealed by molecular simulations.
  Nat Commun, 1, 117.  
19695261 C.C.Su, F.Yang, F.Long, D.Reyon, M.D.Routh, D.W.Kuo, A.K.Mokhtari, J.D.Van Ornam, K.L.Rabe, J.A.Hoy, Y.J.Lee, K.R.Rajashankar, and E.W.Yu (2009).
Crystal structure of the membrane fusion protein CusB from Escherichia coli.
  J Mol Biol, 393, 342-355.
PDB codes: 3h94 3h9i 3h9t 3ooc 3opo 3ow7
19602147 J.Scherer, and D.H.Nies (2009).
CzcP is a novel efflux system contributing to transition metal resistance in Cupriavidus metallidurans CH34.
  Mol Microbiol, 73, 601-621.  
19416927 K.M.Pos (2009).
Trinity revealed: Stoichiometric complex assembly of a bacterial multidrug efflux pump.
  Proc Natl Acad Sci U S A, 106, 6893-6894.  
19453279 T.Eicher, L.Brandstätter, and K.M.Pos (2009).
Structural and functional aspects of the multidrug efflux pump AcrB.
  Biol Chem, 390, 693-699.  
19678712 X.Z.Li, and H.Nikaido (2009).
Efflux-mediated drug resistance in bacteria: an update.
  Drugs, 69, 1555-1623.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.