Escherichia coli K-12 substr. MG1655 Transporter: lipopolysaccharide ABC transporter

Gene: msbA Accession Numbers: EG10613 (EcoCyc), b0914, ECK0905

Regulation Summary Diagram: ?

Regulation summary diagram for msbA

Subunit composition of lipopolysaccharide ABC transporter = [MsbA]2
         ATP-binding lipopolysaccharide transport protein = MsbA

Lipopolysaccharide (LPS), which is a major component of the outer leaflet of the outer membranes of Gram-negative bacteria, consists of lipid A linked to a short core oligosaccharide (lipid A-core) plus a distal O-antigen polysaccharide chain (which is absent in E.coli K-12). MsbA, a member of the ATP Binding Cassette (ABC) Superfamily of transporters [Wu95], is essential for the translocation of the lipid A-core from the inner leaflet to the outer leaflet of the inner membrane [Polissi96, Zhou98b, Doerrler01]. MsbA has also been shown to transport N-acetyl-glucosamine, a precursor of LPS [Polissi96].

The structure of the homodimeric MsbA lipopolysaccharide ABC transporter has been examined using site-directed spin labeling electron paramagnetic resonance spectroscopy [Buchaklian04, Buchaklian05, Dong05, Buchaklian06], and using molecular dynamics simulations and static structural analysis [Campbell03b, Haubertin06]. MsbA is a membrane protein that also possesses a cytoplasmic ATP-binding domain [Polissi96]. Its ability to bind ATP has been demonstrated with an ATP-agarose column [Polissi96]. The structure of MsbA reported by [Chang01] has been retracted [Chang06a]. The crystal structure of MsbA in complex with ADP-vanadate and Ra lipopolysaccharide has been determined to a resolution of 4.2 Å [Reyes05] and in complex with AMPPNP to 3.7 Å resolution [Ward07].

Studies using spheroblasts found that lipopolysaccharides synthesized de novo co-fractionated with the outer membranes in a MsbA-dependent fashion and appeared to be transferred from the inner to outer membrane through contact points between them [Tefsen04]. Newly synthesized lipid A accumulates in the inner leaflet of the inner membrane at the non-permissive temperature in MsbAA270T mutant showing MsbA is required for lipid A flip-flop [Doerrler04, Wang04b]. Experiments measuring lipid flip-flop across the inner membrane suggest MsbA is not required for flip-flop of phospholipids [Kol03].

msbA is a multicopy suppressor of a lpxL mutant and is an essential gene in E. coli [Karow93]. In multicopy, MsbA relieves accumulation of tetra-acylated lipid A and glycerophospholipids in the inner membrane to allow growth of these lpxL mutants [Zhou98b]. msbA-deficient strains were shown to cause the accumulation of hexa-acylated lipid A species and glycerophospholipids within the inner membrane [Zhou98b]. A MsbAA270T mutant is deficient for lipid transport to the outer membrane at the non-permissive temperature [Doerrler01]. A MsbAP18S or MsbAP50S mutant is able to suppress a Kdo deletion phenotype that results in the inability to synthesize Kdo [Mamat08]. The msbA mutants also suppress the lethal waaA deletion mutant which is unable to attach Kdo to lipid IVA [Mamat08]. Multicopy MsbA also partially suppressed a kdsA [Fujishima02] and a kdsD gutQ mutant [Meredith06]. A YhjDR134C mutation makes MsbA dispensable for transport of lipid IVA across the inner membrane and periplasm to the outer membrane [Mamat08]. A msbA mutant showed N-acetyl-[3H]-glucosamine accumulation in the inner membrane, and translocation of N-acetyl-[3H]-glucosamine to the outer membrane was restored by transformation with a plasmid containing the wild-type msbA gene [Polissi96].

The activity of MsbA as a multidrug transporter has been examined by heterologous expression in a lipopolysaccharide-less lactococcal model where expression of MsbA conferred increased resistance to erythromycin, ethidium, and Hoechst 33342 [Woebking05]. Mutational studies indicate that transmembrane helix (TMH) six of MsbA is important in drug binding and transport, specifically of ethidium and Taxol [Woebking08]. MsbA was shown to confer ethidium resistance in an energy-dependent manner, to bind azidopine, and to be stimulated or inhibited by a number of different drugs [Reuter03, Eckford08]. Purified MsbA was also shown to directly eject Hoechst 33342 from the membrane into proteoliposomes (analogous to the periplasm) [Reuter03]. A model for the binding and export of daunorubicin by MsbA has been proposed [Smriti09]. MsbA shares sequence similarity with Mdr1 a human multidrug resistance protein [Karow93].

Review: [Raetz07]

Citations: [Reyes05a, Doerrler07, Borbat07, Miller06, Davidson05a, Shilling03, Zgurskaya02, Schmitt02, Thomas01, Higgins01, Siarheyeva09, Schultz11, Eckford10]

Locations: inner membrane

Map Position: [965,844 -> 967,592] (20.82 centisomes, 75°)
Length: 1749 bp / 582 aa

Molecular Weight of Polypeptide: 64.461 kD (from nucleotide sequence), 65.0 kD (experimental) [Doerrler02 ]

Unification Links: ASAP:ABE-0003113 , CGSC:31743 , DIP:DIP-36229N , DisProt:DP00400 , EchoBASE:EB0608 , EcoGene:EG10613 , EcoliWiki:b0914 , Mint:MINT-1277336 , ModBase:P60752 , OU-Microarray:b0914 , PortEco:msbA , PR:PRO_000023294 , Pride:P60752 , Protein Model Portal:P60752 , RefSeq:NP_415434 , RegulonDB:EG10613 , SMR:P60752 , String:511145.b0914 , UniProt:P60752

Relationship Links: InterPro:IN-FAMILY:IPR003439 , InterPro:IN-FAMILY:IPR003593 , InterPro:IN-FAMILY:IPR011527 , InterPro:IN-FAMILY:IPR011917 , InterPro:IN-FAMILY:IPR017871 , InterPro:IN-FAMILY:IPR027417 , PDB:Structure:3B5W , Pfam:IN-FAMILY:PF00005 , Pfam:IN-FAMILY:PF00664 , Prosite:IN-FAMILY:PS00211 , Prosite:IN-FAMILY:PS50893 , Prosite:IN-FAMILY:PS50929 , Prosite:IN-FAMILY:PS51239 , Smart:IN-FAMILY:SM00382

In Paralogous Gene Group: 23 (75 members)

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0006869 - lipid transport Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA06, Doerrler01]
GO:1901264 - carbohydrate derivative transport Inferred from experiment [Doerrler02]
GO:0006810 - transport Inferred by computational analysis [UniProtGOA11a]
GO:0008152 - metabolic process Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0055085 - transmembrane transport Inferred by computational analysis [GOA01a]
Molecular Function: GO:0005319 - lipid transporter activity Inferred from experiment Inferred by computational analysis [GOA06, Doerrler01]
GO:0005524 - ATP binding Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01a, Polissi96]
GO:0008144 - drug binding Inferred from experiment [Eckford08]
GO:0008289 - lipid binding Inferred from experiment [Eckford08]
GO:0015437 - lipopolysaccharide-transporting ATPase activity Inferred from experiment [Doerrler02]
GO:0034040 - lipid-transporting ATPase activity Inferred from experiment Inferred by computational analysis [GOA01a, Eckford08]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a]
GO:0016887 - ATPase activity Inferred by computational analysis [GOA01a]
GO:0042626 - ATPase activity, coupled to transmembrane movement of substances Inferred by computational analysis [GOA01a]
Cellular Component: GO:0005886 - plasma membrane Inferred from experiment Inferred by computational analysis [UniProtGOA11, UniProtGOA11a, DiazMejia09, Zhang07, Daley05, LopezCampistrou05]
GO:0016020 - membrane Inferred from experiment Inferred by computational analysis [UniProtGOA11a, Lasserre06]
GO:0016021 - integral component of membrane Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01a]
GO:0043190 - ATP-binding cassette (ABC) transporter complex Inferred by computational analysis [GOA06]

MultiFun Terms: cell structure membrane
transport Channel-type Transporters Pyrophosphate Bond (ATP; GTP; P2) Hydrolysis-driven Active Transporters The ATP-binding Cassette (ABC) Superfamily + ABC-type Uptake Permeases ABC superfamily, ATP binding and membrane component

Essentiality data for msbA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB Lennox No 37 Aerobic 7   No [Baba06, Comment 1]

Created 29-May-2008 by Johnson A , JCVI

Enzymatic reaction of: transport of lipid A-core (lipopolysaccharide ABC transporter)

Transport reaction diagram for transport of lipid A-core

Enzymatic reaction of: transport of a lipopolysaccharide (lipopolysaccharide ABC transporter)

EC Number:

Transport reaction diagram for transport of a lipopolysaccharide

Alternative Products for a lipopolysaccharide: hoechst 33342 [Reuter03 ] , ethidium [Reuter03 ]

Cofactors or Prosthetic Groups: Mg2+ [Doerrler02]

Inhibitors (Unknown Mechanism): aluminium fluoride [Eckford08] , beryllium fluoride [Eckford08] , vanadate [Doerrler02]

Sequence Features

Protein sequence of ATP-binding lipopolysaccharide transport protein with features indicated

Feature Class Location Common Name Citations Comment
Transmembrane-Region 25 -> 57 MsbA TM1
Conserved-Region 27 -> 310  
UniProt: ABC transmembrane type-1;
Transmembrane-Region 62 -> 110 MsbA TM2
Transmembrane-Region 63 -> 83  
UniProt: Helical;; Non-Experimental Qualifier: potential;
Transmembrane-Region 121 -> 166 MsbA TM3
Transmembrane-Region 140 -> 162  
UniProt: Helical;; Non-Experimental Qualifier: potential;
Transmembrane-Region 167 -> 186  
UniProt: Helical;; Non-Experimental Qualifier: potential;
Transmembrane-Region 169 -> 209 MsbA TM4
Transmembrane-Region 223 -> 272 MsbA TM5
Transmembrane-Region 253 -> 273  
UniProt: Helical;; Non-Experimental Qualifier: potential;
Mutagenesis-Variant 270  
[Doerrler01, UniProt11]
UniProt: Loss of lipid export.
Transmembrane-Region 275 -> 295  
UniProt: Helical;; Non-Experimental Qualifier: potential;
Transmembrane-Region 280 -> 323 MsbA TM6
Conserved-Region 342 -> 578  
UniProt: ABC transporter;
Nucleotide-Phosphate-Binding-Region 376 -> 383  
UniProt: ATP; Non-Experimental Qualifier: by similarity;
Mutagenesis-Variant 511  
[Polissi96, UniProt10, Schultz11a]
UniProt: Loss of ATPase activity; ATP is still bound;
Mutagenesis-Variant 512  
[Polissi96, Schultz11a]
Increased ATP hydrolysis; decreased cell viability

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Unit:

Transcription-unit diagram


10/20/97 Gene b0914 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10613; confirmed by SwissProt match.


Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Borbat07: Borbat PP, Surendhran K, Bortolus M, Zou P, Freed JH, Mchaourab HS (2007). "Conformational motion of the ABC transporter MsbA induced by ATP hydrolysis." PLoS Biol 5(10);e271. PMID: 17927448

Buchaklian04: Buchaklian AH, Funk AL, Klug CS (2004). "Resting state conformation of the MsbA homodimer as studied by site-directed spin labeling." Biochemistry 43(26);8600-6. PMID: 15222771

Buchaklian05: Buchaklian AH, Klug CS (2005). "Characterization of the Walker A motif of MsbA using site-directed spin labeling electron paramagnetic resonance spectroscopy." Biochemistry 44(14);5503-9. PMID: 15807544

Buchaklian06: Buchaklian AH, Klug CS (2006). "Characterization of the LSGGQ and H motifs from the Escherichia coli lipid A transporter MsbA." Biochemistry 45(41);12539-46. PMID: 17029409

Campbell03b: Campbell JD, Biggin PC, Baaden M, Sansom MS (2003). "Extending the structure of an ABC transporter to atomic resolution: modeling and simulation studies of MsbA." Biochemistry 42(13);3666-73. PMID: 12667056

Chang01: Chang G, Roth CB (2001). "Structure of MsbA from E. coli: a homolog of the multidrug resistance ATP binding cassette (ABC) transporters." Science 293(5536);1793-800. PMID: 11546864

Chang06a: Chang G, Roth CB, Reyes CL, Pornillos O, Chen YJ, Chen AP (2006). "Retraction." Science 314(5807);1875. PMID: 17185584

Daley05: Daley DO, Rapp M, Granseth E, Melen K, Drew D, von Heijne G (2005). "Global topology analysis of the Escherichia coli inner membrane proteome." Science 308(5726);1321-3. PMID: 15919996

Davidson05a: Davidson AL, Chen J (2005). "Structural biology. Flipping lipids: is the third time the charm?." Science 308(5724);963-5. PMID: 15890866

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Doerrler01: Doerrler WT, Reedy MC, Raetz CR (2001). "An Escherichia coli mutant defective in lipid export." J Biol Chem 276(15);11461-4. PMID: 11278265

Doerrler02: Doerrler WT, Raetz CR (2002). "ATPase activity of the MsbA lipid flippase of Escherichia coli." J Biol Chem 277(39);36697-705. PMID: 12119303

Doerrler04: Doerrler WT, Gibbons HS, Raetz CR (2004). "MsbA-dependent translocation of lipids across the inner membrane of Escherichia coli." J Biol Chem 279(43);45102-9. PMID: 15304478

Doerrler07: Doerrler WT (2007). "Density gradient enrichment of Escherichia coli conditional msbA mutants." Appl Environ Microbiol 73(24);7992-6. PMID: 17965211

Dong05: Dong J, Yang G, McHaourab HS (2005). "Structural basis of energy transduction in the transport cycle of MsbA." Science 308(5724);1023-8. PMID: 15890883

Eckford08: Eckford PD, Sharom FJ (2008). "Functional characterization of Escherichia coli MsbA: interaction with nucleotides and substrates." J Biol Chem 283(19);12840-50. PMID: 18344567

Eckford10: Eckford PD, Sharom FJ (2010). "The reconstituted Escherichia coli MsbA protein displays lipid flippase activity." Biochem J 429(1);195-203. PMID: 20412049

Fujishima02: Fujishima H, Nishimura A, Wachi M, Takagi H, Hirasawa T, Teraoka H, Nishimori K, Kawabata T, Nishikawa K, Nagai K (2002). "kdsA mutations affect FtsZ-ring formation in Escherichia coli K-12." Microbiology 148(Pt 1);103-12. PMID: 11782503

GOA01a: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Haubertin06: Haubertin DY, Madaoui H, Sanson A, Guerois R, Orlowski S (2006). "Molecular dynamics simulations of E. coli MsbA transmembrane domain: formation of a semipore structure." Biophys J 91(7);2517-31. PMID: 16782794

Higgins01: Higgins CF, Linton KJ (2001). "Structural biology. The xyz of ABC transporters." Science 293(5536);1782-4. PMID: 11546861

Karow93: Karow M, Georgopoulos C (1993). "The essential Escherichia coli msbA gene, a multicopy suppressor of null mutations in the htrB gene, is related to the universally conserved family of ATP-dependent translocators." Mol Microbiol 7(1);69-79. PMID: 8094880

Kol03: Kol MA, van Dalen A, de Kroon AI, de Kruijff B (2003). "Translocation of phospholipids is facilitated by a subset of membrane-spanning proteins of the bacterial cytoplasmic membrane." J Biol Chem 278(27);24586-93. PMID: 12714595

Lasserre06: Lasserre JP, Beyne E, Pyndiah S, Lapaillerie D, Claverol S, Bonneu M (2006). "A complexomic study of Escherichia coli using two-dimensional blue native/SDS polyacrylamide gel electrophoresis." Electrophoresis 27(16);3306-21. PMID: 16858726

LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532

Mamat08: Mamat U, Meredith TC, Aggarwal P, Kuhl A, Kirchhoff P, Lindner B, Hanuszkiewicz A, Sun J, Holst O, Woodard RW (2008). "Single amino acid substitutions in either YhjD or MsbA confer viability to 3-deoxy-d-manno-oct-2-ulosonic acid-depleted Escherichia coli." Mol Microbiol 67(3);633-48. PMID: 18093093

Meredith06: Meredith TC, Aggarwal P, Mamat U, Lindner B, Woodard RW (2006). "Redefining the requisite lipopolysaccharide structure in Escherichia coli." ACS Chem Biol 1(1);33-42. PMID: 17163638

Miller06: Miller G (2006). "Scientific publishing. A scientist's nightmare: software problem leads to five retractions." Science 314(5807);1856-7. PMID: 17185570

Polissi96: Polissi A, Georgopoulos C (1996). "Mutational analysis and properties of the msbA gene of Escherichia coli, coding for an essential ABC family transporter." Mol Microbiol 1996;20(6);1221-33. PMID: 8809774

Raetz07: Raetz CR, Reynolds CM, Trent MS, Bishop RE (2007). "Lipid A modification systems in gram-negative bacteria." Annu Rev Biochem 76;295-329. PMID: 17362200

Reuter03: Reuter G, Janvilisri T, Venter H, Shahi S, Balakrishnan L, van Veen HW (2003). "The ATP binding cassette multidrug transporter LmrA and lipid transporter MsbA have overlapping substrate specificities." J Biol Chem 278(37);35193-8. PMID: 12842882

Reyes05: Reyes CL, Chang G (2005). "Lipopolysaccharide stabilizes the crystal packing of the ABC transporter MsbA." Acta Crystallograph Sect F Struct Biol Cryst Commun 61(Pt 7);655-8. PMID: 16511120

Reyes05a: Reyes CL, Chang G (2005). "Structure of the ABC transporter MsbA in complex with ADP.vanadate and lipopolysaccharide." Science 308(5724);1028-31. PMID: 15890884

Schmitt02: Schmitt L (2002). "The first view of an ABC transporter: the X-ray crystal structure of MsbA from E. coli." Chembiochem 3(2-3);161-5. PMID: 11921393

Schultz11: Schultz KM, Merten JA, Klug CS (2011). "Characterization of the E506Q and H537A Dysfunctional Mutants in the E. coli ABC Transporter MsbA." Biochemistry 50(18);3599-608. PMID: 21462989

Schultz11a: Schultz KM, Merten JA, Klug CS (2011). "Effects of the L511P and D512G mutations on the Escherichia coli ABC transporter MsbA." Biochemistry 50(13);2594-602. PMID: 21344946

Shilling03: Shilling RA, Balakrishnan L, Shahi S, Venter H, van Veen HW (2003). "A new dimer interface for an ABC transporter." Int J Antimicrob Agents 22(3);200-4. PMID: 13678821

Siarheyeva09: Siarheyeva A, Sharom FJ (2009). "The ABC transporter MsbA interacts with lipid A and amphipathic drugs at different sites." Biochem J 419(2);317-28. PMID: 19132955

Smriti09: Smriti , Zou P, McHaourab HS (2009). "Mapping Daunorubicin-binding Sites in the ATP-binding Cassette Transporter MsbA Using Site-specific Quenching by Spin Labels." J Biol Chem 284(20);13904-13. PMID: 19278995

Tefsen04: Tefsen B, Geurtsen J, Beckers F, Tommassen J, de Cock H (2004). "Lipopolysaccharide transport to the bacterial outer membrane in spheroplasts." J Biol Chem. PMID: 15576375

Thomas01: Thomas PJ, Hunt JF (2001). "A snapshot of Nature's favorite pump." Nat Struct Biol 8(11);920-3. PMID: 11685233

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProt11: UniProt Consortium (2011). "UniProt version 2011-06 released on 2011-06-30 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

Wang04b: Wang X, Karbarz MJ, McGrath SC, Cotter RJ, Raetz CR (2004). "MsbA transporter-dependent lipid A 1-dephosphorylation on the periplasmic surface of the inner membrane: topography of francisella novicida LpxE expressed in Escherichia coli." J Biol Chem 279(47);49470-8. PMID: 15339914

Ward07: Ward A, Reyes CL, Yu J, Roth CB, Chang G (2007). "Flexibility in the ABC transporter MsbA: Alternating access with a twist." Proc Natl Acad Sci U S A 104(48);19005-10. PMID: 18024585

Woebking05: Woebking B, Reuter G, Shilling RA, Velamakanni S, Shahi S, Venter H, Balakrishnan L, van Veen HW (2005). "Drug-lipid A interactions on the Escherichia coli ABC transporter MsbA." J Bacteriol 187(18);6363-9. PMID: 16159769

Woebking08: Woebking B, Velamakanni S, Federici L, Seeger MA, Murakami S, van Veen HW (2008). "Functional role of transmembrane helix 6 in drug binding and transport by the ABC transporter MsbA." Biochemistry 47(41);10904-14. PMID: 18803398

Wu95: Wu LF, Mandrand-Berthelot MA (1995). "A family of homologous substrate-binding proteins with a broad range of substrate specificity and dissimilar biological functions." Biochimie 1995;77(9);744-50. PMID: 8789466

Zgurskaya02: Zgurskaya HI (2002). "Molecular analysis of efflux pump-based antibiotic resistance." Int J Med Microbiol 292(2);95-105. PMID: 12195740

Zhang07: Zhang N, Chen R, Young N, Wishart D, Winter P, Weiner JH, Li L (2007). "Comparison of SDS- and methanol-assisted protein solubilization and digestion methods for Escherichia coli membrane proteome analysis by 2-D LC-MS/MS." Proteomics 7(4);484-93. PMID: 17309111

Zhou98b: Zhou Z, White KA, Polissi A, Georgopoulos C, Raetz CR (1998). "Function of Escherichia coli MsbA, an essential ABC family transporter, in lipid A and phospholipid biosynthesis." J Biol Chem 1998;273(20);12466-75. PMID: 9575204

Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
Page generated by SRI International Pathway Tools version 19.0 on Tue Oct 6, 2015, biocyc12.