Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 37
  Caroff M, Karibian D "Structure of bacterial lipopolysaccharides" - Carbohydrate Research 338(23) (2003) 2431-2447
  

  Bacterial lipopolysaccharides are the major components of the outer surface of Gram-negative bacteria They are often of interest in medicine for their immunomodulatory properties. In small amounts they can be beneficial, but in larger amounts they may cause endotoxic shock. Although they share a common architecture, their structural details exert a strong influence on their activity. These molecules comprise: a lipid moiety, called lipid A, which is considered to be the endotoxic component, a glycosidic part consisting of a core of approximately 10 monosaccharides and, in 'smooth-type' lipopolysaccharides, a third region, named O-chain, consisting of repetitive subunits of one to eight monosaccharides responsible for much of the immunospecificity of the bacterial cell.

  Lipopolysaccharide, structure, core, lipid A, endotoxin, O-chains

  NCBI PubMed ID: 14670707
  Publication DOI: 10.1016/j.carres.2003.07.010
  Journal NLM ID: 0043535
  Publisher: Elsevier
  Correspondence: martine.carloff@bbmpc.u-psud.fr
  Institutions: Equipe Endotoxines, UMR 8619 du Centre National de la Recherche Scientifique, IBBMC, Université de Paris-Sud, F-Orsay, France
  
   
  
  Rhizobium etli
  CSDB ID 31472 (all data & tools)
• Article ID: 1438
  Basu SS, Karbarz MJ, Raetz CR "Expression cloning and characterization of the C28 acyltransferase of lipid A biosynthesis in Rhizobium leguminosarum" - Journal of Biological Chemistry 277(32) (2002) 28959-28971
  

  An unusual feature of lipid A from plant endosymbionts of the Rhizobiaceae family is the presence of a 27-hydroxyoctacosanoic acid (C28) moiety. An enzyme that incorporates this acyl chain is present in extracts of Rhizobium leguminosarum, Rhizobium etli, and Sinorhizobium meliloti but not Escherichia coli. The enzyme transfers 27- hydroxyoctacosanate from a specialized acyl carrier protein (AcpXL) to the precursor Kdo2 ((3-deoxy-d-manno-octulosonic acid)2)-lipid IV(A). We now report the identification of five hybrid cosmids that direct the overexpression of this activity by screening approximately 4000 lysates of individual colonies of an R. leguminosarum 3841 genomic DNA library in the host strain S. meliloti 1021. In these heterologous constructs, both the C28 acyltransferase and C28-AcpXL are overproduced. Sequencing of a 9-kb insert from cosmid pSSB-1, which is also present in the other cosmids, shows that acpXL and the lipid A acyltransferase gene (lpxXL) are close to each other but not contiguous. Nine other open reading frames around lpxXL were also sequenced. Four of them encode orthologues of fatty acid and/or polyketide biosynthetic enzymes. AcpXL purified from S. meliloti expressing pSSB-1 is fully acylated, mainly with 27-hydroxyoctacosanoate. Expression of lpxXL in E. coli behind a T7 promoter results in overproduction in vitro of the expected R. leguminosarum acyltransferase, which is C28-AcpXL-dependent and utilizes (3-deoxy-d-manno-octulosonic acid)2-lipid IV(A) as the acceptor. These findings confirm that lpxXL is the structural gene for the C28 acyltransferase. LpxXL is distantly related to the lauroyltransferase (LpxL) of E. coli lipid A biosynthesis, but highly significant LpxXL orthologues are present in Agrobacterium tumefaciens, Brucella melitensis, and all sequenced strains of Rhizobium, consistent with the occurrence of long secondary acyl chains in the lipid A molecules of these organisms

  biosynthesis, structure, cloning, lipid A, sequencing, Rhizobium leguminosarum, acyltransferase

  NCBI PubMed ID: 12019272
  Publication DOI: 10.1074/jbc.M204525200
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Correspondence: raetz@biochem.duke.edu
  Institutions: Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA
  Methods: SDS-PAGE, DNA techniques, acid hydrolysis, genetic methods, biochemical methods, HPLC, enzyme assay, FPLC, bioinformatic analysis
  
   
  
  Rhizobium etli CE3
  CSDB ID 31643 (all data & tools)
• Article ID: 1477
  Que NLS, Ribeiro AA, Raetz CRH "Two-dimensional NMR spectroscopy and structures of six lipid species from Rhizobium etli. Detection of an acyloxyacyl residue in each component and origin of the aminogluconate moiety" - Journal of Biological Chemistry 275(36) (2001) 28017-28027
  

  The chemical structures of six lipid A species (A, B, C, D-1, D-2, and E) purified from Rhizobium etli CE3 were investigated by one- and two-dimensional NMR spectroscopy. The R. etli lipid A subtypes each contain an unusual acyloxyacyl residue at position 2' as part of a conserved distal glucosamine moiety but differ in their proximal units. All R. etli lipid A species lack phosphate groups. However, they are derivatized with an α-linked galacturonic acid group at position 4', as shown by nuclear Overhauser effect spectroscopy. Component B, which had been not been reported in previous studies, features a β1'-6 linked disaccharide of glucosamine acylated at positions 2, 3, 2', and 3' in a pattern that is typical of lipid A found in other Gram-negative bacteria. D-1 contains an acylated aminogluconate unit in place of the proximal glucosamine residue of B. C and E lack ester-linked beta-hydroxyacyl chains at position 3, as judged by their H-3 chemical shifts, and may be synthesized from B and D-1, respectively, by the R. etli 3-O-deacylase. D-2 is an isomer of D-1 that forms nonenzymatically by acyl chain migration. A may be an elimination product derived from D-1 during hydrolysis at 100 degrees C (pH 4.5), a step needed to release lipid A from lipopolysaccharide. Based on these findings, we propose a biosynthetic scheme for R. etli lipid A in which B is generated first by a variation of the E. coli pathway. The aminogluconate unit of D-1 could then be made from B by enzymatic oxidation of the proximal glucosamine. As predicted by our hypothesis, enzyme(s) can be demonstrated in extracts of R. etli that convert (14)C-labeled B to D-1.

  NMR, structure, Bacterial, lipid A, NMR spectroscopy, Rhizobium, Rhizobium etli, spectroscopy, two-dimensional, acyloxyacyl

  NCBI PubMed ID: 10856304
  Publication DOI: 10.1074/jbc.M004009200
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Correspondence: raetz@biochem.duke.edu
  Institutions: Department of Biochemistry and the Duke NMR Spectroscopy Center and Department of Radiology, Duke University Medical Center, Durham, North Carolina 27710, USA
  Methods: 13C NMR, 1H NMR, NMR-2D, chemical analysis, TLC, MALDI-TOF MS, NMR-1D, DEAE, methylation analysis
  
   
  
  Rhizobium etli CE3, Rhizobium leguminosarum 3855
  CSDB ID 32043 (all data & tools)
• Article ID: 4621
  Sohlenkamp C, Raetz CR, Ingram BO "The calcium-stimulated lipid A 3-O deacylase from Rhizobium etli is not essential for plant nodulation" - Biochimica et Biophysica Acta 1831(7) (2013) 1250-1259
  

  The lipid A component of lipopolysaccharide from the nitrogen-fixing plant endosymbiont, Rhizobium etli, is structurally very different from that found in most enteric bacteria. The lipid A from free-living R. etli is structurally heterogeneous and exists as a mixture of species which are either pentaacylated or tetraacylated. In contrast, the lipid A from R. etli bacteroids is reported to consist exclusively of tetraacylated lipid A species. The tetraacylated lipid A species in both cases lack a β-hydroxymyristoyl chain at the 3-position of lipid A. Here, we show that the lipid A modification enzyme responsible for 3-O deacylation in R. etli is a homolog of the PagL protein originally described in Salmonella enterica sv. typhimurium. In contrast to the PagL proteins described from other species, R. etli PagL displays a calcium dependency. To determine the importance of the lipid A modification catalyzed by PagL, we isolated and characterized a R. etli mutant deficient in the pagL gene. Mass spectrometric analysis confirmed that the mutant strain was exclusively tetraacylated and radiochemical analysis revealed that 3-O deacylase activity was absent in membranes prepared from the mutant. The R. etli mutant was not impaired in its ability to form nitrogen-fixing nodules on Phaseolus vulgaris but it displayed slower nodulation kinetics relative to the wild-type strain. The lipid A modification catalyzed by R. etli PagL, therefore, is not required for nodulation but may play other roles such as protecting bacterial endosymbionts from plant immune responses during infection.

  Lipopolysaccharide, lipid A, Rhizobium etli, Gram-negative bacteria, pagL

  NCBI PubMed ID: 24046865
  Publication DOI: 10.1016/j.bbalip.2013.04.002
  Journal NLM ID: 0217513
  Publisher: Elsevier
  Correspondence: chsohlen@ccg.unam.mx; B.O. Ingram
  Institutions: Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA, Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Av. Universidad s/n, Apdo. Postal 565-A, Cuernavaca, Morelos, CP62210, Mexico
  Methods: DNA techniques, TLC, ESI-MS, biological assays, genetic methods, biochemical methods, LC-MS, assay of polymyxin sensitivity, radiochemical analysis
  
   
  
  Rhizobium etli
  CSDB ID 32041 (all data & tools)

Show legend Show as text

Structure type: oligomer ; 2001.4
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 156
  Vedam V, Kannenberg EL, Haynes JG, Sherrier DJ, Datta A, Carlson RW "A Rhizobium leguminosarum acpXL mutant produces lipopolysaccharide lacking 27-hydroxyoctacosanoic acid" - Journal of Bacteriology 185(6) (2003) 1841-1850
  

  The structure of the lipid A from Rhizobium etli and Rhizobium leguminosarum lipopolysaccharides (LPSs) lacks phosphate and contains a galacturonosyl residue at its 4' position, an acylated 2-aminogluconate in place of the proximal glucosamine, and a very long chain omega-1 hydroxy fatty acid, 27-hydroxyoctacosanoic acid (27OHC28:0). The 27OHC28:0 moiety is common in lipid A's among members of the Rhizobiaceae and also among a number of the facultative intracellular pathogens that form chronic infections, e.g., Brucella abortus, Bartonella henselae, and Legionella pneumophila. In this paper, a mutant of R. leguminosarum was created by placing a kanamycin resistance cassette within acpXL, the gene which encodes the acyl carrier protein for 27OHC28:0. The result was an LPS containing a tetraacylated lipid A lacking 27OHC28:0. A small amount of the mutant lipid A may contain an added palmitic acid residue. The mutant is sensitive to changes in osmolarity and an increase in acidity, growth conditions that likely occur in the nodule microenvironment. In spite of the probably hostile microenvironment of the nodule, the acpXL mutant is still able to form nitrogen-fixing root nodules even though the appearance and development of nodules are delayed. Therefore, it is possible that the acpXL mutant has a host-inducible mechanism which enables it to adapt to these physiological changes.

  Lipopolysaccharide, lipid A, Rhizobium etli, Rhizobium leguminosarum

  NCBI PubMed ID: 12618448
  Publication DOI: 10.1128/JB.185.6.1841-1850.2003
  Journal NLM ID: 2985120R
  Publisher: American Society for Microbiology
  Correspondence: rcarlson@ccrc.uga.edu
  Institutions: Carbohydrate Research Center, University of Georgia Complex, Athens, GA, USA, Department of Microbiology and Biotechnology, University of Tubingen, D072076 Tubingen, Germany, Department of Plant and Soil Sciences, University of Delaware, Newark, Delaware 19717
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, GC, MALDI-TOF MS, composition analysis, genetic methods
  
   
  
  Rhizobium etli, Rhizobium leguminosarum bv. viciae 3841
  CSDB ID 31367 (all data & tools)

Show legend Show as text

Structure type: oligomer ; 1493.2 [M-H]-
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 156
  Vedam V, Kannenberg EL, Haynes JG, Sherrier DJ, Datta A, Carlson RW "A Rhizobium leguminosarum acpXL mutant produces lipopolysaccharide lacking 27-hydroxyoctacosanoic acid" - Journal of Bacteriology 185(6) (2003) 1841-1850
  

  The structure of the lipid A from Rhizobium etli and Rhizobium leguminosarum lipopolysaccharides (LPSs) lacks phosphate and contains a galacturonosyl residue at its 4' position, an acylated 2-aminogluconate in place of the proximal glucosamine, and a very long chain omega-1 hydroxy fatty acid, 27-hydroxyoctacosanoic acid (27OHC28:0). The 27OHC28:0 moiety is common in lipid A's among members of the Rhizobiaceae and also among a number of the facultative intracellular pathogens that form chronic infections, e.g., Brucella abortus, Bartonella henselae, and Legionella pneumophila. In this paper, a mutant of R. leguminosarum was created by placing a kanamycin resistance cassette within acpXL, the gene which encodes the acyl carrier protein for 27OHC28:0. The result was an LPS containing a tetraacylated lipid A lacking 27OHC28:0. A small amount of the mutant lipid A may contain an added palmitic acid residue. The mutant is sensitive to changes in osmolarity and an increase in acidity, growth conditions that likely occur in the nodule microenvironment. In spite of the probably hostile microenvironment of the nodule, the acpXL mutant is still able to form nitrogen-fixing root nodules even though the appearance and development of nodules are delayed. Therefore, it is possible that the acpXL mutant has a host-inducible mechanism which enables it to adapt to these physiological changes.

  Lipopolysaccharide, lipid A, Rhizobium etli, Rhizobium leguminosarum

  NCBI PubMed ID: 12618448
  Publication DOI: 10.1128/JB.185.6.1841-1850.2003
  Journal NLM ID: 2985120R
  Publisher: American Society for Microbiology
  Correspondence: rcarlson@ccrc.uga.edu
  Institutions: Carbohydrate Research Center, University of Georgia Complex, Athens, GA, USA, Department of Microbiology and Biotechnology, University of Tubingen, D072076 Tubingen, Germany, Department of Plant and Soil Sciences, University of Delaware, Newark, Delaware 19717
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, GC, MALDI-TOF MS, composition analysis, genetic methods
  
   
  
  Rhizobium leguminosarum bv. viciae 22
  CSDB ID 31369 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 699
  Jeyaretnam B, Glushka J, Kolli VS, Carlson RW "Characterization of a novel lipid-A from Rhizobium species Sin-1. A UNIQUE LIPID-A STRUCTURE THAT IS DEVOID OF PHOSPHATE AND HAS A GLYCOSYL BACKBONE CONSISTING OF GLUCOSAMINE AND 2-AMINOGLUCONIC ACID" - Journal of Biological Chemistry 277(44) (2002) 41802-41810
  

  The structure of the lipid-A from Rhizobium species Sin-1, a nitrogen-fixing Gram-negative bacterial symbiont of Sesbania, was determined by composition, nuclear magnetic resonance spectroscopic, and mass spectrometric analyses. The lipid-A preparation consisted of a mixture of structures due to differences in fatty acylation and in the glycosyl backbone. There were two different disaccharide backbones. One disaccharide consisted of a distal glucosaminosyl residue β-linked to position 6 of a proximal 2-aminoglucono-1,5-lactonosyl residue, and in the second disaccharide, the proximal residue was 2-amino-2,3- dideoxy-d-erythro-hex-2-enono-1,5-lactone. For both disaccharides, the distal glucosaminewas acylated at C-2' primarily with b-hydroxypalmitate (b-OHC16:0) which, in turn, was O-acylated with 27-hydroxyoctacosanoic acid. For some of the lipid-A molecules, the distal glucosaminosyl residue was also acylated at C-3' with b-hydroxymyristate (b-OHC14:0), whereas other molecules were devoid of this acyl substituent. Both the 2-aminoglucono-1,5-lactonosyl and 2-amino-2,3-dideoxy-D-erythro-hex-2-enono-1,5-lactonosyl residues were acylated at C-2, primarily with b-OHC16:0. Minor amounts of lipid-A molecules contained b-OHC14:0 at C-3 and/or b-hydroxystearate (b-OHC18:0) or b-hydroxyoctadecenoate (b-OHC18:1) asthe C-2 and C-2' N-acyl substituents.

  NMR, structure, characterization, lipid A, Rhizobium, disaccharide, species, nitrogen fixing

  NCBI PubMed ID: 12193590
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Correspondence: RCARLSON@ccrc.uga.edu
  Institutions: Complex Carbohydrate Research Center, The University of Georgia, Athens, Georgia, USA
  Methods: NMR
  
   
  
  Rhizobium sp. Sin-1
  CSDB ID 942 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 699
  Jeyaretnam B, Glushka J, Kolli VS, Carlson RW "Characterization of a novel lipid-A from Rhizobium species Sin-1. A UNIQUE LIPID-A STRUCTURE THAT IS DEVOID OF PHOSPHATE AND HAS A GLYCOSYL BACKBONE CONSISTING OF GLUCOSAMINE AND 2-AMINOGLUCONIC ACID" - Journal of Biological Chemistry 277(44) (2002) 41802-41810
  

  The structure of the lipid-A from Rhizobium species Sin-1, a nitrogen-fixing Gram-negative bacterial symbiont of Sesbania, was determined by composition, nuclear magnetic resonance spectroscopic, and mass spectrometric analyses. The lipid-A preparation consisted of a mixture of structures due to differences in fatty acylation and in the glycosyl backbone. There were two different disaccharide backbones. One disaccharide consisted of a distal glucosaminosyl residue β-linked to position 6 of a proximal 2-aminoglucono-1,5-lactonosyl residue, and in the second disaccharide, the proximal residue was 2-amino-2,3- dideoxy-d-erythro-hex-2-enono-1,5-lactone. For both disaccharides, the distal glucosaminewas acylated at C-2' primarily with b-hydroxypalmitate (b-OHC16:0) which, in turn, was O-acylated with 27-hydroxyoctacosanoic acid. For some of the lipid-A molecules, the distal glucosaminosyl residue was also acylated at C-3' with b-hydroxymyristate (b-OHC14:0), whereas other molecules were devoid of this acyl substituent. Both the 2-aminoglucono-1,5-lactonosyl and 2-amino-2,3-dideoxy-D-erythro-hex-2-enono-1,5-lactonosyl residues were acylated at C-2, primarily with b-OHC16:0. Minor amounts of lipid-A molecules contained b-OHC14:0 at C-3 and/or b-hydroxystearate (b-OHC18:0) or b-hydroxyoctadecenoate (b-OHC18:1) asthe C-2 and C-2' N-acyl substituents.

  NMR, structure, characterization, lipid A, Rhizobium, disaccharide, species, nitrogen fixing

  NCBI PubMed ID: 12193590
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Correspondence: RCARLSON@ccrc.uga.edu
  Institutions: Complex Carbohydrate Research Center, The University of Georgia, Athens, Georgia, USA
  Methods: NMR
  
   
  
  Rhizobium sp. Sin-1
  CSDB ID 804 (all data & tools)
• Article ID: 4988
  Choma A, Komaniecka I, Zebracki K "Structure, biosynthesis and function of unusual lipids A from nodule-inducing and N2-fixing bacteria" - Biochimica et Biophysica Acta 1862(2) (2017) 196-209
  

  This review focuses on the chemistry and structures of (Brady)rhizobium lipids A, indispensable parts of lipopolysaccharides. These lipids contain unusual (omega-1) hydroxylated very long chain fatty acids, which are synthesized by a very limited group of bacteria, besides rhizobia. The significance and requirement of the very long chain fatty acids for outer membrane stability as well as the genetics of the synthesis pathway are discussed. The biological role of these fatty acids for bacterial life in extremely different environments (soil and intracellular space within nodules) is also considered.

  lipid A, Rhizobium, Nodule-forming bacteria, Very long chain fatty acids (VLCFA)

  NCBI PubMed ID: 27836696
  Publication DOI: 10.1016/j.bbalip.2016.11.004
  Journal NLM ID: 0217513
  Publisher: Elsevier
  Correspondence: adam.choma@poczta.umcs.lublin.pl
  Institutions: Department of Genetics and Microbiology, Maria Curie-Sklodowska University, Akademicka 19, 20-033 Lublin, Poland
  
   
  
  Rhizobium sp. Sin-1, Rhizobium galegae
  CSDB ID 12070 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 1439
  Gazzano-Santoro H, Parent JB, Conlon PJ, Kasler HG, Tsai C, Lill-Elghanian DA, Hollingsworth RI "Characterization of the structural elements in lipid A required for binding of a recombinant fragment of bactericidal/permeability-increasing protein rBPI23" - Infection and Immunity 63 (1995) 2201-2205
  

  Both human bactericidal/permeability-increasing protein (BPI) and a recombinant amino-terminal fragment of BPI (rBPI23) have been shown to bind with high affinity to the lipid A region of lipopolysaccharide (LPS) (H. Gazzano-Santoro, J. B. Parent, L. Grinna, A. Horwitz, T. Parsons, G. Theofan, P. Elsbach, J. Weiss, and P. J. Conlon, Infect. Immun. 60:4754-4761, 1992). In the present study, lipid A preparations derived from bacterial LPS as well as synthetic lipid A's and various lipid A analogs were used to determine the structural elements required for rBPI23 binding. rBPI23 bound in vitro to a variety of synthetic and natural lipid A preparations (both mono- and diphosphoryl forms), including lipid A's prepared from Escherichia coli and Salmonella, Neisseria, and Rhizobium species. Binding does not require that the origin of negative charge be phosphate, since rBPI23 bound with high affinity to lipid A's isolated from Rhizobium species that contain carboxylate (Rhizobium trifolii) or sulfate (Rhizobium meliloti) anionic groups and lack phosphate. Lipid A acyl chains are important, since rBPI23 did not bind to four synthetic variants of the β(1-6)-linked D-glucosamine disaccharide lipid A head group, all devoid of acyl chains. rBPI23 also bound weakly to lipid X, a monosaccharide lipid precursor of LPS corresponding to the reducing half of lipid A. Lipid IVA, a precursor identical to E. coli lipid A except that it lacks the 2' and 3' acyl chains, was the simplest structure identified in this study that rBPI23 bound with high affinity. These results demonstrate that rBPI23 has a binding specificity for the lipid A region of LPS and binding involves both electrostatic and hydrophobic components.

  structural, characterization, lipid, lipid A, protein, fragment, binding, recombinant, bactericidal/permeability increasing protein

  NCBI PubMed ID: 7768599
  Journal NLM ID: 0246127
  WWW link: http://iai.asm.org/content/63/6/2201.full.pdf
  Publisher: American Society for Microbiology
  Institutions: Sepsis Research Department, XOMA Corporation, Berkeley, California 947101, Neurocrine Biosciences, San Diego, California 921212, Department of Health and Human Services, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD, USA3, and Departments of Biochemistry and Chemistry, Michigan State University, East Lansing, Michigan 48824
  Methods: radiolabeling, serological methods
  
   
  
  Rhizobium leguminosarum bv. trifolii
  CSDB ID 31308 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: glycolipid
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 2677
  Bhat UR, Forsberg LS, Carlson RW "Structure of lipid A component of Rhizobium leguminosarum bv. phaseoli lipopolysaccharide. Unique nonphosphorylated lipid A containing 2-amino-2-deoxygluconate, galacturonate, and glucosamine" - Journal of Biological Chemistry 269 (1994) 14402-14410
  

  The structure of lipid A from the lipopolysaccharide of Rhizobium leguminosarum bv. phaseoli (wild type strain CE3) was investigated by alkylation analysis, nuclear magnetic resonance spectroscopy, and electrospray and fast atom bombardment mass spectrometry of the de-O-acylated lipid A. The lipid A carbohydrate backbone was shown to be a trisaccharide containing galacturonic acid, glucosamine, and the unique sugar 2-amino-2-deoxygluconic acid, previously unreported in lipopolysaccharides. Nuclear magnetic resonance spectroscopy and ethylation analyses revealed that the galacturonic acid is α-1,4-linked to the glucosamine, while the amino aldonic acid residue, which may exist as the 1,5-lactone, is attached as an aglycone to the glucosamine and, thus, occupies the reducing end of the molecule. The resulting backbone is hydrophilic and analogous to the commonly observed bisphosphorylated glucosamine disaccharide from enteric bacterial lipopolysaccharides in that both the nonreducing and reducing ends carry negatively charged substituents. The fatty acids of the R. leguminosarum lipid A are attached both as O- and N-acyl substituents to glucosamine and 2-aminogluconate. All fatty acids are hydroxylated consisting of 3-hydroxymyristate (3-OH-C14.0), 3-hydroxypentadecanoate (3-OH-C15.0), 3-hydroxypalmitate (3-OH-C16.0), 3-hydroxystearate (3-OH-C18.0), and 27-hydroxyoctacosanoate (27-OH-C28.0) in the approximate mole ratio 3:0.2:1:0.6:1. Unlike lipid As from enteric bacteria, the R. leguminosarum lipid A lacks 3-acyloxyacyl substituents; however, the long chain 27-hydroxy fatty acid carries ester-linked β-hydroxybutyrate at the 27-hydroxy position. Fast atom bombardment mass spectrometry of the de-O-acylated lipid A demonstrated the presence of 2 molecular species that differ by 28 mass units due to fatty acid heterogeneity at the two amide linkages. One species carries amide-linked 3-OH-C14.0 and 3-OH-C16.0; the second species carries 3-OH-C14.0 and 3-OH-C18.0. Each molecular species also exists as the aldonolactone, yielding molecular ions at ((M+H)+)-18. The heterogeneity in the amide-linked fatty acids further distinguishes the Rhizobium lipid A from enteric lipid As.

  NCBI PubMed ID: 8182046
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Institutions: Complex Carbohydrate Research Center, University of Georgia, Athens 30605
  
   
  
  Rhizobium leguminosarum bv. phaseoli
  CSDB ID 137109 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: LPS
Contained glycoepitopes: IEDB_115136,IEDB_130650,IEDB_130659,IEDB_130699,IEDB_130701,IEDB_136045,IEDB_136906,IEDB_137472,IEDB_140630,IEDB_141794,IEDB_141807,IEDB_142489,IEDB_144562,IEDB_144983,IEDB_151528,IEDB_151531,IEDB_152206,IEDB_152214,IEDB_174333,IEDB_190606,IEDB_983930,SB_44,SB_67,SB_7,SB_72,SB_86

• Article ID: 3416
  D'Haeze W, Leoff C, Freshour G, Noel KD, Carlson RW "Rhizobium etli CE3 bacteroid lipopolysaccharides are structurally similar but not identical to those produced by cultured CE3 bacteria" - Journal of Biological Chemistry 282(23) (2007) 17101-17113
  

  Rhizobium etli CE3 bacteroids were isolated from Phaseolus vulgaris root nodules. The LPS from the bacteroids was purified and compared with the LPS from laboratory-cultured R. etli CE3 and from cultures grown in the presence of anthocyanin. Comparisons were made of the O-chain polysaccharide, the core oligosaccharide, and the lipid A. It was found that, while LPS from CE3 bacteria and bacteroids are structurally similar, bacteroid LPS had specific modifications to both the O-chain polysaccharide and lipid A portions of their LPS. Cultures grown with anthocyanin contained modifications only to the O-chain polysaccharide. The changes to the O-chain polysaccharide consisted of the addition of a single methyl group to the 2-position of a fucosyl residue in one of the five O-chain trisaccharide repeat units. This same change occurred for bacteria grown in the presence of anthocyanin. This methylation change correlated with the inability of bacteroid LPS and LPS from anthocyanin-containing cultures to bind the monoclonal antibody, JIM28. The core oligosaccharide region of bacteroid LPS and from anthocyanin-grown cultures was identical to that of LPS from normal laboratory-cultured CE3. The lipid A from bacteroids consisted exclusively of a tetraacylated species compared to the presence of both tetra- and pentaacylated lipid A from laboratory cultures. Growth in the presence of anthocyanin did not affect the lipid A structure. Purified bacteroids that could resume growth were also found to be more sensitive to the cationic peptides, poly-L-lysine, polymyxin-B, and melittin

  Lipopolysaccharide, lipid A, monoclonal antibodies, Rhizobium etli, modification, root nodules

  NCBI PubMed ID: 17420254
  Publication DOI: 10.1074/jbc.M611669200
  Journal NLM ID: 2985121R
  Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
  Correspondence: rcarlson@ccrc.uga.edu
  Institutions: Complex Carbohydrate Research Center, The University of Georgia, Athens, GA 30602
  Methods: GC-MS, mild acid hydrolysis, MALDI-TOF MS, composition analysis, serological methods, electron microscopy, immunoblotting
  
   
  
  Rhizobium etli CE3
  CSDB ID 22048 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 3794
  Vanderlinde EM, Muszynski A, Harrison JJ, Koval SF, Foreman DL, Ceri H, Kannenberg EL, Carlson RW, Yost CK "Rhizobium leguminosarum biovar viciae 3841, deficient in 27-hydroxyoctacosanoate-modified lipopolysaccharide, is impaired in desiccation tolerance, biofilm formation and motility." - Microbiology (2009) 3055-3069
  

  The lipopolysaccharide (LPS) of the Gram-negative legume symbiont Rhizobium leguminosarum biovar viciae 3841 contains several unique modifications, including the addition of a 27-hydroxyoctacosanoic acid (27OHC28:0), also termed the very long chain fatty acid (VLCFA), attached at the 2' position of lipid A. A transposon mutant that lacks expression of two putative 3-oxo-acyl [acyl-carrier protein] synthase II genes, fabF1 and fabF2, from the VLCFA biosynthetic cluster, was isolated and characterized. Mass spectrometry indicated that the lipid A of the mutant lacked the VLCFA modification and DOC-PAGE of the LPS indicated further structural alterations. The mutant was characteristically sensitive to several stresses that would be experienced in the soil environment, such as desiccation and osmotic stresses. An increase in the excretion of neutral surface polysaccharides was observed in the mutant. This mutant was also altered in its attachment to solid surfaces, and was non-motile, with most of the mutant cells lacking flagella. Despite the pleiotropic effects of the mutation, these mutants were still able to nodulate legumes and fix atmospheric nitrogen. This report emphasizes that a structurally intact VLCFA containing lipid A is critical to cellular traits that are important for survival in the rhizosphere.

  Lipopolysaccharide, gene, lipid A, mass spectrometry, modification, formation, Rhizobium leguminosarum, Biofilm, motility

  NCBI PubMed ID: 19460825
  Publication DOI: 10.1099/mic.0.025031-0
  Journal NLM ID: 0376646
  Publisher: Washington, DC: Kluwer Academic/Plenum Publishers
  Correspondence: chris.yost@uregina.ca
  Institutions: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA, Department of Biology, University of Regina, 3737 Wascana Parkway, Regina, SK S4S 0A2, Canada, Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada, Department of Microbiology and Immunology, University of Western Ontario, London, ON N6A 5C1, Canada
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, composition analysis, genetic methods, RT-PCR, microscopy, MALDI-FOF MS
  
   
  
  Rhizobium leguminosarum bv. viciae 3841
  CSDB ID 31221 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 3794
  Vanderlinde EM, Muszynski A, Harrison JJ, Koval SF, Foreman DL, Ceri H, Kannenberg EL, Carlson RW, Yost CK "Rhizobium leguminosarum biovar viciae 3841, deficient in 27-hydroxyoctacosanoate-modified lipopolysaccharide, is impaired in desiccation tolerance, biofilm formation and motility." - Microbiology (2009) 3055-3069
  

  The lipopolysaccharide (LPS) of the Gram-negative legume symbiont Rhizobium leguminosarum biovar viciae 3841 contains several unique modifications, including the addition of a 27-hydroxyoctacosanoic acid (27OHC28:0), also termed the very long chain fatty acid (VLCFA), attached at the 2' position of lipid A. A transposon mutant that lacks expression of two putative 3-oxo-acyl [acyl-carrier protein] synthase II genes, fabF1 and fabF2, from the VLCFA biosynthetic cluster, was isolated and characterized. Mass spectrometry indicated that the lipid A of the mutant lacked the VLCFA modification and DOC-PAGE of the LPS indicated further structural alterations. The mutant was characteristically sensitive to several stresses that would be experienced in the soil environment, such as desiccation and osmotic stresses. An increase in the excretion of neutral surface polysaccharides was observed in the mutant. This mutant was also altered in its attachment to solid surfaces, and was non-motile, with most of the mutant cells lacking flagella. Despite the pleiotropic effects of the mutation, these mutants were still able to nodulate legumes and fix atmospheric nitrogen. This report emphasizes that a structurally intact VLCFA containing lipid A is critical to cellular traits that are important for survival in the rhizosphere.

  Lipopolysaccharide, gene, lipid A, mass spectrometry, modification, formation, Rhizobium leguminosarum, Biofilm, motility

  NCBI PubMed ID: 19460825
  Publication DOI: 10.1099/mic.0.025031-0
  Journal NLM ID: 0376646
  Publisher: Washington, DC: Kluwer Academic/Plenum Publishers
  Correspondence: chris.yost@uregina.ca
  Institutions: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA, Department of Biology, University of Regina, 3737 Wascana Parkway, Regina, SK S4S 0A2, Canada, Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada, Department of Microbiology and Immunology, University of Western Ontario, London, ON N6A 5C1, Canada
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, composition analysis, genetic methods, RT-PCR, microscopy, MALDI-FOF MS
  
   
  
  Rhizobium leguminosarum bv. viciae 3841
  CSDB ID 31222 (all data & tools)
• Article ID: 4988
  Choma A, Komaniecka I, Zebracki K "Structure, biosynthesis and function of unusual lipids A from nodule-inducing and N2-fixing bacteria" - Biochimica et Biophysica Acta 1862(2) (2017) 196-209
  

  This review focuses on the chemistry and structures of (Brady)rhizobium lipids A, indispensable parts of lipopolysaccharides. These lipids contain unusual (omega-1) hydroxylated very long chain fatty acids, which are synthesized by a very limited group of bacteria, besides rhizobia. The significance and requirement of the very long chain fatty acids for outer membrane stability as well as the genetics of the synthesis pathway are discussed. The biological role of these fatty acids for bacterial life in extremely different environments (soil and intracellular space within nodules) is also considered.

  lipid A, Rhizobium, Nodule-forming bacteria, Very long chain fatty acids (VLCFA)

  NCBI PubMed ID: 27836696
  Publication DOI: 10.1016/j.bbalip.2016.11.004
  Journal NLM ID: 0217513
  Publisher: Elsevier
  Correspondence: adam.choma@poczta.umcs.lublin.pl
  Institutions: Department of Genetics and Microbiology, Maria Curie-Sklodowska University, Akademicka 19, 20-033 Lublin, Poland
  
   
  
  Rhizobium etli CE3, Rhizobium leguminosarum bv. trifolii, Rhizobium leguminosarum bv. viciae 3841
  CSDB ID 12069 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 3794
  Vanderlinde EM, Muszynski A, Harrison JJ, Koval SF, Foreman DL, Ceri H, Kannenberg EL, Carlson RW, Yost CK "Rhizobium leguminosarum biovar viciae 3841, deficient in 27-hydroxyoctacosanoate-modified lipopolysaccharide, is impaired in desiccation tolerance, biofilm formation and motility." - Microbiology (2009) 3055-3069
  

  The lipopolysaccharide (LPS) of the Gram-negative legume symbiont Rhizobium leguminosarum biovar viciae 3841 contains several unique modifications, including the addition of a 27-hydroxyoctacosanoic acid (27OHC28:0), also termed the very long chain fatty acid (VLCFA), attached at the 2' position of lipid A. A transposon mutant that lacks expression of two putative 3-oxo-acyl [acyl-carrier protein] synthase II genes, fabF1 and fabF2, from the VLCFA biosynthetic cluster, was isolated and characterized. Mass spectrometry indicated that the lipid A of the mutant lacked the VLCFA modification and DOC-PAGE of the LPS indicated further structural alterations. The mutant was characteristically sensitive to several stresses that would be experienced in the soil environment, such as desiccation and osmotic stresses. An increase in the excretion of neutral surface polysaccharides was observed in the mutant. This mutant was also altered in its attachment to solid surfaces, and was non-motile, with most of the mutant cells lacking flagella. Despite the pleiotropic effects of the mutation, these mutants were still able to nodulate legumes and fix atmospheric nitrogen. This report emphasizes that a structurally intact VLCFA containing lipid A is critical to cellular traits that are important for survival in the rhizosphere.

  Lipopolysaccharide, gene, lipid A, mass spectrometry, modification, formation, Rhizobium leguminosarum, Biofilm, motility

  NCBI PubMed ID: 19460825
  Publication DOI: 10.1099/mic.0.025031-0
  Journal NLM ID: 0376646
  Publisher: Washington, DC: Kluwer Academic/Plenum Publishers
  Correspondence: chris.yost@uregina.ca
  Institutions: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA, Department of Biology, University of Regina, 3737 Wascana Parkway, Regina, SK S4S 0A2, Canada, Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada, Department of Microbiology and Immunology, University of Western Ontario, London, ON N6A 5C1, Canada
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, composition analysis, genetic methods, RT-PCR, microscopy, MALDI-FOF MS
  
   
  
  Rhizobium leguminosarum bv. viciae fabF2/F1, Rhizobium leguminosarum bv. viciae fabF2
  CSDB ID 31225 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531,IEDB_534865

• Article ID: 3794
  Vanderlinde EM, Muszynski A, Harrison JJ, Koval SF, Foreman DL, Ceri H, Kannenberg EL, Carlson RW, Yost CK "Rhizobium leguminosarum biovar viciae 3841, deficient in 27-hydroxyoctacosanoate-modified lipopolysaccharide, is impaired in desiccation tolerance, biofilm formation and motility." - Microbiology (2009) 3055-3069
  

  The lipopolysaccharide (LPS) of the Gram-negative legume symbiont Rhizobium leguminosarum biovar viciae 3841 contains several unique modifications, including the addition of a 27-hydroxyoctacosanoic acid (27OHC28:0), also termed the very long chain fatty acid (VLCFA), attached at the 2' position of lipid A. A transposon mutant that lacks expression of two putative 3-oxo-acyl [acyl-carrier protein] synthase II genes, fabF1 and fabF2, from the VLCFA biosynthetic cluster, was isolated and characterized. Mass spectrometry indicated that the lipid A of the mutant lacked the VLCFA modification and DOC-PAGE of the LPS indicated further structural alterations. The mutant was characteristically sensitive to several stresses that would be experienced in the soil environment, such as desiccation and osmotic stresses. An increase in the excretion of neutral surface polysaccharides was observed in the mutant. This mutant was also altered in its attachment to solid surfaces, and was non-motile, with most of the mutant cells lacking flagella. Despite the pleiotropic effects of the mutation, these mutants were still able to nodulate legumes and fix atmospheric nitrogen. This report emphasizes that a structurally intact VLCFA containing lipid A is critical to cellular traits that are important for survival in the rhizosphere.

  Lipopolysaccharide, gene, lipid A, mass spectrometry, modification, formation, Rhizobium leguminosarum, Biofilm, motility

  NCBI PubMed ID: 19460825
  Publication DOI: 10.1099/mic.0.025031-0
  Journal NLM ID: 0376646
  Publisher: Washington, DC: Kluwer Academic/Plenum Publishers
  Correspondence: chris.yost@uregina.ca
  Institutions: Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA 30602, USA, Department of Biology, University of Regina, 3737 Wascana Parkway, Regina, SK S4S 0A2, Canada, Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada, Department of Microbiology and Immunology, University of Western Ontario, London, ON N6A 5C1, Canada
  Methods: GC-MS, mild acid hydrolysis, DOC-PAGE, composition analysis, genetic methods, RT-PCR, microscopy, MALDI-FOF MS
  
   
  
  Rhizobium leguminosarum bv. viciae fabF2/F1, Rhizobium leguminosarum bv. viciae fabF2
  CSDB ID 31227 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141181,IEDB_141807,IEDB_151531,IEDB_534865

• Article ID: 3821
  Banoub JH, El Aneed A, Cohen AM, Joly N "Structural investigation of bacterial lipopolysaccharides by mass spectrometry and tandem mass spectrometry" - Mass Spectrometry Reviews 29(4) (2010) 606-650
  

  Mass spectrometric studies are now playing a leading role in the elucidation of lipopolysaccharide (LPS) structures through the characterization of antigenic polysaccharides, core oligosaccharides and lipid A components including LPS genetic modifications. The conventional MS and MS/MS analyses together with CID fragmentation provide additional structural information complementary to the previous analytical experiments, and thus contribute to an integrated strategy for the simultaneous characterization and correct sequencing of the carbohydrate moiety.

  LPS, O-antigen, lipid A, core oligosaccharide, MS and MS/MS analyses

  NCBI PubMed ID: 20589944
  Publication DOI: 10.1002/mas.20258
  Journal NLM ID: 8219702
  Publisher: Wiley
  Correspondence: joe.banoub@dfo-mpo.gc.ca
  Institutions: Fisheries and Oceans Canada, Science Branch, Special Projects, P.O. Box 5667, St. John's, Newfoundland, Canada A1C 5X1, Department of Chemistry, Memorial University of Newfoundland, St. John's, Newfoundland, Canada A1B 3V6, College of Pharmacy and Nutrition, University of Saskatchewan, Thorvaldson Building, 110 Science Place, Saskatoon, Saskatchewan, Canada S7N 5C9, Unité de Catalyse et de Chimie du Solide, Site de l'Artois—UMR CNRS 8181, I.U.T. de Béthune, Département Chimie, 1230 rue de l'Université, BP819, 62408 Béthune Cedex, France, Institute for Marine Biosciences Room 219A, (NRC-IMB), National Research Council of Canada, Government of Canada, 1411 Oxford Street, Halifax, NS, Canada B3H 3Z1
  Methods: MS/MS, MS
  
   
  
  Rhizobium etli
  CSDB ID 31124 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 3863
  Ingram BO, Sohlenkamp C, Geiger O, Raetz CR "Altered lipid A structures and polymyxin hypersensitivity of Rhizobium etli mutants lacking the LpxE and LpxF phosphatases" - Biochimica et Biophysica Acta 1801(5) (2010) 593-604
  

  The lipid A of Rhizobium etli, a nitrogen-fixing plant endosymbiont, displays significant structural differences when compared to that of Escherichia coli. An especially striking feature of R. etli lipid A is that it lacks both the 1- and 4'-phosphate groups. The 4'-phosphate moiety of the distal glucosamine unit is replaced with a galacturonic acid residue. The dephosphorylated proximal unit is present as a mixture of the glucosamine hemiacetal and an oxidized 2-aminogluconate derivative. Distinct lipid A phosphatases directed to the 1 or the 4'-positions have been identified previously in extracts of R. etli and Rhizobium leguminosarum. The corresponding structural genes, lpxE and lpxF, respectively, have also been identified. Here, we describe the isolation and characterization of R. etli deletion mutants in each of these phosphatase genes and the construction of a double phosphatase mutant. Mass spectrometry confirmed that the mutant strains completely lacked the wild-type lipid A species and accumulated the expected phosphate-containing derivatives. Moreover, radiochemical analysis revealed that phosphatase activity was absent in membranes prepared from the mutants. Our results indicate that LpxE and LpxF are solely responsible for selectively dephosphorylating the lipid A molecules of R. etli. All the mutant strains showed an increased sensitivity to polymyxin relative to the wild-type. However, despite the presence of altered lipid A species containing one or both phosphate groups, all the phosphatase mutants formed nitrogen-fixing nodules on Phaseolus vulgaris. Therefore, the dephosphorylation of lipid A molecules in R. etli is not required for nodulation but may instead play a role in protecting the bacteria from cationic antimicrobial peptides or other immune responses of plants.

  Lipopolysaccharide, Rhizobium etli, Gram-negative bacteria, outer membranes, lipid A phosphatases, LpxE, LpxF

  NCBI PubMed ID: 20153447
  Publication DOI: 10.1016/j.bbalip.2010.02.001
  Journal NLM ID: 0217513
  Publisher: Elsevier
  Correspondence: C.R.H. Raetz
  Institutions: Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA, Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Av. Universidad s/n, Apdo. Postal 565-A, Cuernavaca, Morelos, CP62210, Mexico
  Methods: DNA techniques, TLC, ESI-MS, mild acid hydrolysis, biological assays, genetic methods, biochemical methods, assay of polymyxin sensitivity
  
   
  
  Rhizobium etli
  CSDB ID 31199 (all data & tools)

Show legend Show as text

Structure type: oligomer
Compound class: lipid A
Contained glycoepitopes: IEDB_141807,IEDB_151531

• Article ID: 3863
  Ingram BO, Sohlenkamp C, Geiger O, Raetz CR "Altered lipid A structures and polymyxin hypersensitivity of Rhizobium etli mutants lacking the LpxE and LpxF phosphatases" - Biochimica et Biophysica Acta 1801(5) (2010) 593-604
  

  The lipid A of Rhizobium etli, a nitrogen-fixing plant endosymbiont, displays significant structural differences when compared to that of Escherichia coli. An especially striking feature of R. etli lipid A is that it lacks both the 1- and 4'-phosphate groups. The 4'-phosphate moiety of the distal glucosamine unit is replaced with a galacturonic acid residue. The dephosphorylated proximal unit is present as a mixture of the glucosamine hemiacetal and an oxidized 2-aminogluconate derivative. Distinct lipid A phosphatases directed to the 1 or the 4'-positions have been identified previously in extracts of R. etli and Rhizobium leguminosarum. The corresponding structural genes, lpxE and lpxF, respectively, have also been identified. Here, we describe the isolation and characterization of R. etli deletion mutants in each of these phosphatase genes and the construction of a double phosphatase mutant. Mass spectrometry confirmed that the mutant strains completely lacked the wild-type lipid A species and accumulated the expected phosphate-containing derivatives. Moreover, radiochemical analysis revealed that phosphatase activity was absent in membranes prepared from the mutants. Our results indicate that LpxE and LpxF are solely responsible for selectively dephosphorylating the lipid A molecules of R. etli. All the mutant strains showed an increased sensitivity to polymyxin relative to the wild-type. However, despite the presence of altered lipid A species containing one or both phosphate groups, all the phosphatase mutants formed nitrogen-fixing nodules on Phaseolus vulgaris. Therefore, the dephosphorylation of lipid A molecules in R. etli is not required for nodulation but may instead play a role in protecting the bacteria from cationic antimicrobial peptides or other immune responses of plants.

  Lipopolysaccharide, Rhizobium etli, Gram-negative bacteria, outer membranes, lipid A phosphatases, LpxE, LpxF

  NCBI PubMed ID: 20153447
  Publication DOI: 10.1016/j.bbalip.2010.02.001
  Journal NLM ID: 0217513
  Publisher: Elsevier
  Correspondence: C.R.H. Raetz
  Institutions: Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA, Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, Av. Universidad s/n, Apdo. Postal 565-A, Cuernavaca, Morelos, CP62210, Mexico
  Methods: DNA techniques, TLC, ESI-MS, mild acid hydrolysis, biological assays, genetic methods, biochemical methods, assay of polymyxin sensitivity
  
   
  
  Rhizobium etli
  CSDB ID 31200 (all data & tools)
• Article ID: 3864
  Ingram BO, Masoudi A, Raetz CR "Escherichia coli mutants that synthesize dephosphorylated lipid A molecules" - Biochemistry 49(38) (2010) 8325-8337
  

  The lipid A moiety of Escherichia coli lipopolysaccharide is a hexaacylated disaccharide of glucosamine that is phosphorylated at the 1 and 4' positions. Expression of the Francisella novicida lipid A 1-phosphatase FnLpxE in E. coli results in dephosphorylation of the lipid A proximal unit. Coexpression of FnLpxE and the Rhizobium leguminosarum lipid A oxidase RlLpxQ in E. coli converts much of the proximal glucosamine to 2-amino-2-deoxygluconate. Expression of the F. novicida lipid A 4'-phosphatase FnLpxF in wild-type E. coli has no effect because FnLpxF cannot dephosphorylate hexaacylated lipid A. However, expression of FnLpxF in E. coli lpxM mutants, which synthesize pentaacylated lipid A lacking the secondary 3'-myristate chain, causes extensive 4'-dephosphorylation. Coexpression of FnLpxE and FnLpxF in lpxM mutants results in massive accumulation of lipid A species lacking both phosphate groups, and introduction of RlLpxQ generates phosphate-free lipid A variants containing 2-amino-2-deoxygluconate. The proposed lipid A structures were confirmed by electrospray ionization mass spectrometry. Strains with 4'-dephosphorylated lipid A display increased polymyxin resistance. Heptose-deficient mutants of E. coli lacking both the 1- and 4'-phosphate moieties are viable on plates but sensitive to CaCl(2). Our methods for reengineering lipid A structure may be useful for generating novel vaccines and adjuvants.

  lipopolysaccharides, Escherichia coli, Rhizobia, mass spectrometry, lipid A structure, polymyxin

  NCBI PubMed ID: 20795687
  Journal NLM ID: 0370623
  Publisher: American Chemical Society
  Correspondence: raetz@biochem.duke.edu
  Institutions: Department of Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA
  Methods: DNA techniques, ESI-MS, mild acid hydrolysis, genetic methods, enzymatic modification
  
   
  
  Rhizobium leguminosarum
  CSDB ID 31196 (all data & tools)