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MetaCyc Pathway: superpathway of lipopolysaccharide biosynthesis

If an enzyme name is shown in bold, there is experimental evidence for this enzymatic activity.

Superclasses: Biosynthesis Fatty Acids and Lipids Biosynthesis
Superpathways

Some taxa known to possess this pathway include ? : Escherichia coli K-12 substr. MG1655

Expected Taxonomic Range: Bacteria

Summary:
General Background

Bacterial lipopolysaccharides (LPS) are unique and complex glycolipids that provide characteristic components of the outer monolayer of the outer membranes of gram-negative bacteria. Wild type LPS is composed of a hydrophobic domain lipid A (or endotoxin), a phosphorylated "core" oligosaccharide (core OS) and a distal O-specific polysaccharide (or O-antigen). Strains of Escherichia coli K-12 normally do not make O-antigen, unless a mutation in the O-antigen operon is corrected [Stevenson94]. O antigens are not needed for growth in the laboratory, but they protect bacteria from antibiotics and complement-mediated lysis. The lipid A and Kdo domains of LPS are required for growth [Raetz07, Raetz02].

The lipid A (endotoxin) domain of LPS is a unique, glucosamine-based saccharolipid that serves as the hydrophobic anchor of LPS. It is the bioactive component of the molecule that is associated with gram-negative septic shock [Trent04]. There are approximately 106 lipid A residues in an E. coli cell [Raetz07]. Lipid A of E. coli consists of a β (1'->6)-linked D-glucosamine disaccharide that carries four (R)-3-hydroxytetradecanoyl groups in positions 2, 3, 2', and 3' and two phosphoryl residues in positions 1 and 4'. The hydroxy fatty acids at positions 2' and 3' are acylated at their 3-hydroxyl groups by dodecanoic acid and tetradecanoic acid, respectively [Rietschel]. Given their conserved architecture, most types of lipid A molecules are detected at picomolar levels by TLR4 (toll-like receptor 4) of the innate immune system present on macrophages and endothelial cells of animals [Raetz02].

The core oligosaccharides are conceptually divided into two regions: the inner core (lipid A proximal) and the outer core. The inner core is highly conserved. It comprises three 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and L-glycero-D-manno-heptose (Hep) residues and it is often phosphorylated. Structural diversity in the core oligosaccharide is limited by the constraints imposed by its essential role in outer membrane stability [Whitfield03, Heinrichs98]. E. coli mutants that lack the heptose region of the inner core display the "deep rough" phenotype characterized by changes in both the structure and the composition of the outer membrane leading to its instability [Irvin79]. Deep rough mutants of E. coli activate expression of colanic acid exopolysaccharide, lose expression of pili and flagella and secrete a form of hemolysin with reduced hemolytic activity [Parker92]. The outer core comprises a tri-hexose backbone modified with varying side-branch substitutions of hexose and acetamidohexose residues. The outer core region provides an attachment site for O antigen [Heinrichs98]. Mutations that eliminate synthesis of the outer core result in increased susceptibility to some hydrophobic compounds due to an indirect effect on core phosphorylation [Austin90].

About This Pathway

E. coli lipid A is synthesized on the cytoplasmic surface of the inner membrane by a conserved pathway of nine constitutive enzymes. The first reaction of lipid A biosynthesis is the acylation of the sugar nucleotide UDP-GlcNAc by LpxA. Deacetylation of the product, UDP-3-O-(acyl)-GlcNAc by the zinc metalloenzyme LpxC is the committed reaction of the pathway. LpxC is an excellent target for the design of novel, gram-negative specific antibiotics. Following deacetylation, a second beta-hydroxymyristate moiety is incorporated by LpxD to generate UDP-2,3-diacylglucosamine. UDP-2,3-diacylglucosamine is cleaved at its pyrophosphate bond by the highly selective pyrophosphatase LpxH to form 2,3-diacylglucosamine-1-phosphate (lipid X). A beta,1'-6 linked disaccharide is then generated by the condensation of another molecule of UDP-2,3-diacylglucosamine with lipid X by LpxB. A specific kinase, LpxK next phosphorylates the 4' position of the disaccharide to form lipid IVA. The kinase product, lipid IVA, is of great interest because it possesses some of the properties of endotoxins [Raetz02].

E. coli LPS contains two Kdo residues that are transferred to lipid IVA by a bifunctional enzyme, encoded by waaA. The chromosomal waa region (formerly rfa) contains the major core-oligosaccharide assembly operons, and E. coli K-12 provided the first waa region sequenced in its entirety [Whitfield03]. The last steps of E. coli lipid A biosynthesis involve the addition of lauroyl and myristoyl residues to the distal glucosamine unit (generating acyloxyacyl moieties by LpxL and LpxM respectively. The completed lipid A-Kdo2 serves as the acceptor on which the core oligosaccharide and lipo-oligosaccharide chains are assembled.

The lipid A-core portion (known as rough LPS (R-LPS)) is synthesized by a conserved pathway. The mechanism involves sequential glycosyl transfer from nucleotide sugar precursors with a co-coordinated complex of membrane-associated glycosyltransferases (Waa proteins) acting at the cytoplasmic surface of the inner membrane (IM). The modification of the core region of E. coli LPS requires the action of three enzymes, viz. WaaP (an LPS kinase), WaaY (an enzyme required for a second phosphorylation) and WaaQ (a transferase that adds the side-branch heptose). The WaaP enzyme is the most important of these activities since the modifications must proceed in the strict order WaaPQY. Mutants lacking WaaP activity exhibit the deep rough phenotype and are avirulent. Mutant LPSs lacking the outer core (waaG) are phosphorylated inefficiently by WaaP [Yethon00]. Conversely, the loss of phosphorylation results in a decreased efficiency of core extension, leading to more truncated cores than seen in the wild type.

The lipid A-core is then flipped to the outer surface of the inner membrane by the ATP-binding cassette (ABC) transporter, MsbA. An additional integral membrane protein, YhjD, has recently been implicated in LPS export across the IM. The smallest LPS derivative that supports viability in E. coli is lipid IVA. However, it requires mutations in either MsbA or YhjD, to suppress the normally lethal consequence of an incomplete lipid A [Tran08]. Recent studies with deletion mutants implicate the periplasmic protein LptA, the cytosolic protein LptB, and the IM proteins LptC, LptF, and LptG in the subsequent transport of nascent LPS to the outer membrane (OM), where the LptD/LptE complex flips LPS to the outer surface [Ma08].

This topic has been reviewed in [Raetz07, Trent04, Whitfield03, Raetz02, Heinrichs98, Whitfield97, Rietschel93, Takada89, Rietschel, Jann].

Subpathways: lipid IVA biosynthesis , CMP-KDO biosynthesis I , KDO transfer to lipid IVA I , (KDO)2-lipid A biosynthesis I , Lipid A-core biosynthesis

Unification Links: EcoCyc:LPSSYN-PWY

Credits:
Created 13-Jul-2006 by Johnson A , TIGR
Reviewed 25-Mar-2010 by Sarker M , SRI International


References

Austin90: Austin EA, Graves JF, Hite LA, Parker CT, Schnaitman CA (1990). "Genetic analysis of lipopolysaccharide core biosynthesis by Escherichia coli K-12: insertion mutagenesis of the rfa locus." J Bacteriol 172(9);5312-25. PMID: 2168379

Heinrichs98: Heinrichs DE, Yethon JA, Whitfield C (1998). "Molecular basis for structural diversity in the core regions of the lipopolysaccharides of Escherichia coli and Salmonella enterica." Mol Microbiol 30(2);221-32. PMID: 9791168

Irvin79: Irvin RT, Lam J, Costerton JW (1979). "Structural and biochemical examination of ghosts derived from a deep rough (heptose-deficient lipopolysaccharide) strain and a smooth strain of Escherichia coli." Can J Microbiol 25(4);436-46. PMID: 385128

Jann: Jann K, Jann B "Polysaccharide antigens of Escherichia coli." Rev Infect Dis 9 Suppl 5;S517-26. PMID: 2446369

Ma08: Ma B, Reynolds CM, Raetz CR (2008). "Periplasmic orientation of nascent lipid A in the inner membrane of an Escherichia coli LptA mutant." Proc Natl Acad Sci U S A 105(37);13823-8. PMID: 18768814

Parker92: Parker CT, Kloser AW, Schnaitman CA, Stein MA, Gottesman S, Gibson BW (1992). "Role of the rfaG and rfaP genes in determining the lipopolysaccharide core structure and cell surface properties of Escherichia coli K-12." J Bacteriol 174(8);2525-38. PMID: 1348243

Raetz02: Raetz CR, Whitfield C (2002). "Lipopolysaccharide endotoxins." Annu Rev Biochem 71;635-700. PMID: 12045108

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

Rietschel: Rietschel ET, Brade L, Brandenburg K, Flad HD, de Jong-Leuveninck J, Kawahara K, Lindner B, Loppnow H, Luderitz T, Schade U "Chemical structure and biologic activity of bacterial and synthetic lipid A." Rev Infect Dis 9 Suppl 5;S527-36. PMID: 3317748

Rietschel93: Rietschel ET, Kirikae T, Schade FU, Ulmer AJ, Holst O, Brade H, Schmidt G, Mamat U, Grimmecke HD, Kusumoto S (1993). "The chemical structure of bacterial endotoxin in relation to bioactivity." Immunobiology 187(3-5);169-90. PMID: 8330896

Stevenson94: Stevenson G, Neal B, Liu D, Hobbs M, Packer NH, Batley M, Redmond JW, Lindquist L, Reeves P (1994). "Structure of the O antigen of Escherichia coli K-12 and the sequence of its rfb gene cluster." J Bacteriol 1994;176(13);4144-56. PMID: 7517391

Takada89: Takada H, Kotani S (1989). "Structural requirements of lipid A for endotoxicity and other biological activities." Crit Rev Microbiol 16(6);477-523. PMID: 2663021

Tran08: Tran AX, Trent MS, Whitfield C (2008). "The LptA protein of Escherichia coli is a periplasmic lipid A-binding protein involved in the lipopolysaccharide export pathway." J Biol Chem 283(29);20342-9. PMID: 18480051

Trent04: Trent MS (2004). "Biosynthesis, transport, and modification of lipid A." Biochem Cell Biol 82(1);71-86. PMID: 15052329

Whitfield03: Whitfield C, Kaniuk N, Frirdich E (2003). "Molecular insights into the assembly and diversity of the outer core oligosaccharide in lipopolysaccharides from Escherichia coli and Salmonella." J Endotoxin Res 9(4);244-9. PMID: 12935355

Whitfield97: Whitfield C, Amor PA, Koplin R (1997). "Modulation of the surface architecture of gram-negative bacteria by the action of surface polymer:lipid A-core ligase and by determinants of polymer chain length." Mol Microbiol 23(4);629-38. PMID: 9157235

Yethon00: Yethon JA, Vinogradov E, Perry MB, Whitfield C (2000). "Mutation of the lipopolysaccharide core glycosyltransferase encoded by waaG destabilizes the outer membrane of Escherichia coli by interfering with core phosphorylation." J Bacteriol 182(19);5620-3. PMID: 10986272

Other References Related to Enzymes, Genes, Subpathways, and Substrates of this Pathway

Agladze05: Agladze K, Wang X, Romeo T (2005). "Spatial periodicity of Escherichia coli K-12 biofilm microstructure initiates during a reversible, polar attachment phase of development and requires the polysaccharide adhesin PGA." J Bacteriol 187(24);8237-46. PMID: 16321928

Airoldi10: Airoldi C, Sommaruga S, Merlo S, Sperandeo P, Cipolla L, Polissi A, Nicotra F (2010). "Targeting bacterial membranes: NMR spectroscopy characterization of substrate recognition and binding requirements of D-arabinose-5-phosphate isomerase." Chemistry 16(6);1897-902. PMID: 20039350

Anderson85: Anderson MS, Bulawa CE, Raetz CR (1985). "The biosynthesis of gram-negative endotoxin. Formation of lipid A precursors from UDP-GlcNAc in extracts of Escherichia coli." J Biol Chem 1985;260(29);15536-41. PMID: 3905795

Anderson87: Anderson MS, Raetz CR (1987). "Biosynthesis of lipid A precursors in Escherichia coli. A cytoplasmic acyltransferase that converts UDP-N-acetylglucosamine to UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine." J Biol Chem 262(11);5159-69. PMID: 3549716

Anderson88: Anderson MS, Robertson AD, Macher I, Raetz CR (1988). "Biosynthesis of lipid A in Escherichia coli: identification of UDP-3-O-[(R)-3-hydroxymyristoyl]-alpha-D-glucosamine as a precursor of UDP-N2,O3-bis[(R)-3-hydroxymyristoyl]-alpha-D-glucosamine." Biochemistry 1988;27(6);1908-17. PMID: 3288280

Anderson92: Anderson MS, Raetz CR (1992). "UDP-N-acetylglucosamine 3-O-acyltransferase from Escherichia coli." Methods Enzymol 1992;209;449-54. PMID: 1495425

Anderson93: Anderson MS, Bull HG, Galloway SM, Kelly TM, Mohan S, Radika K, Raetz CR (1993). "UDP-N-acetylglucosamine acyltransferase of Escherichia coli. The first step of endotoxin biosynthesis is thermodynamically unfavorable." J Biol Chem 1993;268(26);19858-65. PMID: 8366124

Armstrong06: Armstrong MT, Theg SM, Braun N, Wainwright N, Pardy RL, Armstrong PB (2006). "Histochemical evidence for lipid A (endotoxin) in eukaryote chloroplasts." FASEB J 20(12);2145-6. PMID: 16935939

Asojo01: Asojo O, Friedman J, Adir N, Belakhov V, Shoham Y, Baasov T (2001). "Crystal structures of KDOP synthase in its binary complexes with the substrate phosphoenolpyruvate and with a mechanism-based inhibitor." Biochemistry 40(21);6326-34. PMID: 11371194

Baasov93: Baasov T, Sheffer-Dee-Noor S, Kohen A, Jakob A, Belakhov V (1993). "Catalytic mechanism of 3-deoxy-D-manno-2-octulosonate-8-phosphate synthase. The use of synthetic analogues to probe the structure of the putative reaction intermediate." Eur J Biochem 217(3);991-9. PMID: 8223657

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

Babinski02: Babinski KJ, Ribeiro AA, Raetz CR (2002). "The Escherichia coli gene encoding the UDP-2,3-diacylglucosamine pyrophosphatase of lipid A biosynthesis." J Biol Chem 277(29);25937-46. PMID: 12000770

Babinski02a: Babinski KJ, Kanjilal SJ, Raetz CR (2002). "Accumulation of the lipid A precursor UDP-2,3-diacylglucosamine in an Escherichia coli mutant lacking the lpxH gene." J Biol Chem 277(29);25947-56. PMID: 12000771

Badger05: Badger J, Sauder JM, Adams JM, Antonysamy S, Bain K, Bergseid MG, Buchanan SG, Buchanan MD, Batiyenko Y, Christopher JA, Emtage S, Eroshkina A, Feil I, Furlong EB, Gajiwala KS, Gao X, He D, Hendle J, Huber A, Hoda K, Kearins P, Kissinger C, Laubert B, Lewis HA, Lin J, Loomis K, Lorimer D, Louie G, Maletic M, Marsh CD, Miller I, Molinari J, Muller-Dieckmann HJ, Newman JM, Noland BW, Pagarigan B, Park F, Peat TS, Post KW, Radojicic S, Ramos A, Romero R, Rutter ME, Sanderson WE, Schwinn KD, Tresser J, Winhoven J, Wright TA, Wu L, Xu J, Harris TJ (2005). "Structural analysis of a set of proteins resulting from a bacterial genomics project." Proteins 60(4);787-96. PMID: 16021622

Barb08: Barb AW, Zhou P (2008). "Mechanism and inhibition of LpxC: an essential zinc-dependent deacetylase of bacterial lipid A synthesis." Curr Pharm Biotechnol 9(1);9-15. PMID: 18289052

Bartling08: Bartling CM, Raetz CR (2008). "Steady-state kinetics and mechanism of LpxD, the N-acyltransferase of lipid A biosynthesis." Biochemistry 47(19);5290-302. PMID: 18422345

Bartling09: Bartling CM, Raetz CR (2009). "Crystal structure and acyl chain selectivity of Escherichia coli LpxD, the N-acyltransferase of lipid A biosynthesis." Biochemistry 48(36);8672-83. PMID: 19655786

Bateman99: Bateman A (1999). "The SIS domain: a phosphosugar-binding domain." Trends Biochem Sci 24(3);94-5. PMID: 10203754

Beall87: Beall B, Lutkenhaus J (1987). "Sequence analysis, transcriptional organization, and insertional mutagenesis of the envA gene of Escherichia coli." J Bacteriol 169(12);5408-15. PMID: 2824434

Beher81: Beher MG, Schnaitman CA (1981). "Regulation of the OmpA outer membrane protein of Escherichia coli." J Bacteriol 147(3);972-85. PMID: 7024253

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Report Errors or Provide Feedback
Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
Page generated by SRI International Pathway Tools version 18.5 on Wed Nov 26, 2014, biocyc11.