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Escherichia coli K-12 substr. MG1655 Enzyme: UDP-3-O-acyl-N-acetylglucosamine deacetylase



Gene: lpxC Accession Numbers: EG10265 (EcoCyc), b0096, ECK0097

Synonyms: asmB, envA

Regulation Summary Diagram: ?

Summary:
UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) catalyzes the second reaction and the first committed step in lipid A biosynthesis. LpxC hydrolyzes UDP-3-O-(3-hydroxymyristoyl)-N-acetlyglucosamine to UDP-3-O-(3-hydroxymyristoyl)glucosamine and acetate [Young95, Anderson88]. The mechanism of action of LpxC has been studied, and a high-throughput fluorescent screen as well as a mass-spectrometry based screen have been developed to evaluate LpxC activity [Hernick05, Wang01e, Langsdorf10].

LpxC is a metalloenzyme that requires bound zinc for activity [Jackman99]. More recently, LpxC with bound iron was shown to have higher activity than zinc as the cofactor [Hernick10]. LpxC has significantly higher affinity for zinc, however, the in vivo concentrations of readily available iron are higher. The ability of LpxC to switch between metal ions in response to availability has been proposed. [Gattis10] SIte directed mutagenesis has identified zinc binding sites [Jackman01].

LpxC abundance is regulated in response to the cell's need for lipid A [Sorensen96a]. This regulation occurs via degradation by HflB, integral membrane ATP-dependent zinc metallopeptidase, and requires the presence of the carboxy-terminus of LpxC which contains a signal sequence necessary for this FtsH-dependent degradation [Ogura99, Fuhrer06]. LpxC is degraded by FtsH in a growth-rate dependent manner, where LpxC is degraded during slow growth, and stabilized when cells grow rapidly [Schakermann13].

lpxC is an essential gene [Beall87]. Mutations in lpxC lead to problems with cell division, increased permeability to antibacterial agents, leakage of periplasmic enzymes and reduced abundance of LPS [Normark69, Normark70, Young91, Kloser96].

LpxC is a drug target for antimicrobials due to its role in lipid A synthesis [Onishi96]. Various inhibitors of LpxC have been discovered [Clements02, Jackman00, Barb08, Liang11a, Liang13, Loppenberg13]. The molecular determinants of binding and specificity as well as the catalytic mechanism have been studied with respect to drug development [Hernick06]. Spontaneous mutants which were resistant to LpxC inhibitors were isolated and found to have mutations in fabZ or thrS [Zeng13].

The crystal structure of LpxC in complex with UDP-(3-O-(R-3-hydroxymyristoyl))-glucosamine was determined [Clayton13]. The crystal structure analysis of LpxC bound to an inhibitor revealed a backbone flipping of a loop causing a conformational switch allowing for expansion of the active site and thereby accomodating LpxC inhibitors with different head groups [Lee14].

Citations: [Langklotz11, Westphal12]

Gene Citations: [Vicente98, MenginLecreulx98, Hara97]

Locations: cytosol

Map Position: [106,557 -> 107,474] (2.3 centisomes)
Length: 918 bp / 305 aa

Molecular Weight of Polypeptide: 33.956 kD (from nucleotide sequence), 33.96 kD (experimental) [Jackman99 ]

pI: 5.87

Unification Links: ASAP:ABE-0000336 , CGSC:815 , DIP:DIP-48045N , EchoBASE:EB0261 , EcoGene:EG10265 , EcoliWiki:b0096 , Mint:MINT-1309717 , ModBase:P0A725 , OU-Microarray:b0096 , PortEco:lpxC , PR:PRO_000023122 , Protein Model Portal:P0A725 , RefSeq:NP_414638 , RegulonDB:EG10265 , SMR:P0A725 , String:511145.b0096 , UniProt:P0A725

Relationship Links: InterPro:IN-FAMILY:IPR004463 , InterPro:IN-FAMILY:IPR011334 , InterPro:IN-FAMILY:IPR015870 , InterPro:IN-FAMILY:IPR020568 , Pfam:IN-FAMILY:PF03331

Gene-Reaction Schematic: ?

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0009245 - lipid A biosynthetic process Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA01a, Young95]
GO:0006629 - lipid metabolic process Inferred by computational analysis [UniProtGOA11a]
Molecular Function: GO:0005506 - iron ion binding Inferred from experiment [Gattis10]
GO:0008270 - zinc ion binding Inferred from experiment [Jackman99]
GO:0008759 - UDP-3-O-[3-hydroxymyristoyl] N-acetylglucosamine deacetylase activity Inferred from experiment Inferred by computational analysis [GOA01a, Jackman99]
GO:0019213 - deacetylase activity Inferred from experiment [Anderson88]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005737 - cytoplasm Inferred from experiment [Anderson88]
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: cell structure surface antigens (ECA, O antigen of LPS)
metabolism biosynthesis of macromolecules (cellular constituents) lipopolysaccharide lipid A

Essentiality data for lpxC knockouts: ?

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

Credits:
Curated 18-Aug-2006 by Shearer A , SRI International
Last-Curated ? 24-Oct-2013 by Kubo A , SRI International


Enzymatic reaction of: UDP-3-O-acyl-N-acetylglucosamine deacetylase

EC Number: 3.5.1.108

UDP-3-O-[(3R)-3-hydroxymyristoyl]-N-acetyl-α-D-glucosamine + H2O <=> UDP-3-O-(3-hydroxymyristoyl)-α-D-glucosamine + acetate

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

The reaction is irreversible in the direction shown.

In Pathways: superpathway of (KDO)2-lipid A biosynthesis , superpathway of lipopolysaccharide biosynthesis , lipid IVA biosynthesis

Inhibitors (Unknown Mechanism): Zn2+ [Jackman99] , EDTA [Jackman99] , dipicolinate [Jackman99] , N-ethylmaleimide [Anderson88]

Primary Physiological Regulators of Enzyme Activity: Zn2+

Kinetic Parameters:

Substrate
Km (μM)
Citations
UDP-3-O-[(3R)-3-hydroxymyristoyl]-N-acetyl-α-D-glucosamine
2.1
[Jackman99]


Sequence Features

Feature Class Location Citations Comment
Extrinsic-Sequence-Variant 50
[UniProt10]
Alternate sequence: F → S; UniProt: (in ASMB2/3; reduced activity);
Metal-Binding-Site 79, 238, 246, 265
These residues make up the zinc-binding site. Histidines 79 and 238 appear to directly coordinate zinc, with additional coordination provided by one of the two other residues. All four are essential for catalytic activity [Jackman01]. Extended X-ray absorption fine structure spectroscopy suggests that aspartate-246 coordinates zinc with the aid of a solvent molecule [McClure03].
Extrinsic-Sequence-Variant 210
[UniProt10]
Alternate sequence: G → S; UniProt: (in ASMB1; reduced activity);


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b0096 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10265; confirmed by SwissProt match.


References

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

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

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

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

Clayton13: Clayton GM, Klein DJ, Rickert KW, Patel SB, Kornienko M, Zugay-Murphy J, Reid JC, Tummala S, Sharma S, Singh SB, Miesel L, Lumb KJ, Soisson SM (2013). "Structure of the bacterial deacetylase LpxC bound to the nucleotide reaction product reveals mechanisms of oxyanion stabilization and proton transfer." J Biol Chem. PMID: 24108127

Clements02: Clements JM, Coignard F, Johnson I, Chandler S, Palan S, Waller A, Wijkmans J, Hunter MG (2002). "Antibacterial activities and characterization of novel inhibitors of LpxC." Antimicrob Agents Chemother 46(6);1793-9. PMID: 12019092

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

Fuhrer06: Fuhrer F, Langklotz S, Narberhaus F (2006). "The C-terminal end of LpxC is required for degradation by the FtsH protease." Mol Microbiol 59(3);1025-36. PMID: 16420369

Gattis10: Gattis SG, Hernick M, Fierke CA (2010). "Active site metal ion in UDP-3-O-((R)-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC) switches between Fe(II) and Zn(II) depending on cellular conditions." J Biol Chem 285(44);33788-96. PMID: 20709752

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

Hara97: Hara H, Yasuda S, Horiuchi K, Park JT (1997). "A promoter for the first nine genes of the Escherichia coli mra cluster of cell division and cell envelope biosynthesis genes, including ftsI and ftsW." J Bacteriol 179(18);5802-11. PMID: 9294438

Hernick05: Hernick M, Gennadios HA, Whittington DA, Rusche KM, Christianson DW, Fierke CA (2005). "UDP-3-O-((R)-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase functions through a general acid-base catalyst pair mechanism." J Biol Chem 280(17);16969-78. PMID: 15705580

Hernick06: Hernick M, Fierke CA (2006). "Catalytic mechanism and molecular recognition of E. coli UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase probed by mutagenesis." Biochemistry 45(51);15240-8. PMID: 17176046

Hernick10: Hernick M, Gattis SG, Penner-Hahn JE, Fierke CA (2010). "Activation of Escherichia coli UDP-3-O-[(R)-3-hydroxymyristoyl]-N-acetylglucosamine deacetylase by Fe2+ yields a more efficient enzyme with altered ligand affinity." Biochemistry 49(10);2246-55. PMID: 20136146

Jackman00: Jackman JE, Fierke CA, Tumey LN, Pirrung M, Uchiyama T, Tahir SH, Hindsgaul O, Raetz CR (2000). "Antibacterial agents that target lipid A biosynthesis in gram-negative bacteria. Inhibition of diverse UDP-3-O-(r-3-hydroxymyristoyl)-n-acetylglucosamine deacetylases by substrate analogs containing zinc binding motifs." J Biol Chem 275(15);11002-9. PMID: 10753902

Jackman01: Jackman JE, Raetz CR, Fierke CA (2001). "Site-directed mutagenesis of the bacterial metalloamidase UDP-(3-O-acyl)-N-acetylglucosamine deacetylase (LpxC). Identification of the zinc binding site." Biochemistry 40(2);514-23. PMID: 11148046

Jackman99: Jackman JE, Raetz CR, Fierke CA (1999). "UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase of Escherichia coli is a zinc metalloenzyme." Biochemistry 1999;38(6);1902-11. PMID: 10026271

Kloser96: Kloser AW, Laird MW, Misra R (1996). "asmB, a suppressor locus for assembly-defective OmpF mutants of Escherichia coli, is allelic to envA (lpxC)." J Bacteriol 178(17);5138-43. PMID: 8752330

Langklotz11: Langklotz S, Schakermann M, Narberhaus F (2011). "Control of lipopolysaccharide biosynthesis by FtsH-mediated proteolysis of LpxC is conserved in enterobacteria but not in all gram-negative bacteria." J Bacteriol 193(5);1090-7. PMID: 21193611

Langsdorf10: Langsdorf EF, Malikzay A, Lamarr WA, Daubaras D, Kravec C, Zhang R, Hart R, Monsma F, Black T, Ozbal CC, Miesel L, Lunn CA (2010). "Screening for antibacterial inhibitors of the UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC) using a high-throughput mass spectrometry assay." J Biomol Screen 15(1);52-61. PMID: 20019290

Lee14: Lee CJ, Liang X, Gopalaswamy R, Najeeb J, Ark ED, Toone EJ, Zhou P (2013). "Structural Basis of the Promiscuous Inhibitor Susceptibility of E. coli LpxC." ACS Chem Biol. PMID: 24117400

Liang11a: Liang X, Lee CJ, Chen X, Chung HS, Zeng D, Raetz CR, Li Y, Zhou P, Toone EJ (2011). "Syntheses, structures and antibiotic activities of LpxC inhibitors based on the diacetylene scaffold." Bioorg Med Chem 19(2);852-60. PMID: 21194954

Liang13: Liang X, Lee CJ, Zhao J, Toone EJ, Zhou P (2013). "Synthesis, structure, and antibiotic activity of aryl-substituted LpxC inhibitors." J Med Chem 56(17);6954-66. PMID: 23914798

Loppenberg13: Loppenberg M, Muller H, Pulina C, Oddo A, Teese M, Jose J, Holl R (2013). "Synthesis and biological evaluation of flexible and conformationally constrained LpxC inhibitors." Org Biomol Chem 11(36);6056-70. PMID: 23917427

McClure03: McClure CP, Rusche KM, Peariso K, Jackman JE, Fierke CA, Penner-Hahn JE (2003). "EXAFS studies of the zinc sites of UDP-(3-O-acyl)-N-acetylglucosamine deacetylase (LpxC)." J Inorg Biochem 94(1-2);78-85. PMID: 12620676

MenginLecreulx98: Mengin-Lecreulx D, Ayala J, Bouhss A, van Heijenoort J, Parquet C, Hara H (1998). "Contribution of the Pmra promoter to expression of genes in the Escherichia coli mra cluster of cell envelope biosynthesis and cell division genes." J Bacteriol 180(17);4406-12. PMID: 9721276

Normark69: Normark S (1969). "Transduction and dominance studies of the envA gene present in a chain-forming mutant of Escherichia coli K 12." J Gen Microbiol 57(3);xxx. PMID: 4903169

Normark70: Normark S (1970). "Genetics of a chain-forming mutant of Escherichia coli. Transduction and dominance of the envA gene mediating increased penetration to some antibacterial agents." Genet Res 16(1);63-78. PMID: 4922970

Ogura99: Ogura T, Inoue K, Tatsuta T, Suzaki T, Karata K, Young K, Su LH, Fierke CA, Jackman JE, Raetz CR, Coleman J, Tomoyasu T, Matsuzawa H (1999). "Balanced biosynthesis of major membrane components through regulated degradation of the committed enzyme of lipid A biosynthesis by the AAA protease FtsH (HflB) in Escherichia coli." Mol Microbiol 31(3);833-44. PMID: 10048027

Onishi96: Onishi HR, Pelak BA, Gerckens LS, Silver LL, Kahan FM, Chen MH, Patchett AA, Galloway SM, Hyland SA, Anderson MS, Raetz CR (1996). "Antibacterial agents that inhibit lipid A biosynthesis." Science 274(5289);980-2. PMID: 8875939

Schakermann13: Schakermann M, Langklotz S, Narberhaus F (2013). "FtsH-mediated coordination of lipopolysaccharide biosynthesis in Escherichia coli correlates with the growth rate and the alarmone (p)ppGpp." J Bacteriol 195(9);1912-9. PMID: 23417489

Sorensen96a: Sorensen PG, Lutkenhaus J, Young K, Eveland SS, Anderson MS, Raetz CR (1996). "Regulation of UDP-3-O-[R-3-hydroxymyristoyl]-N-acetylglucosamine deacetylase in Escherichia coli. The second enzymatic step of lipid a biosynthesis." J Biol Chem 271(42);25898-905. PMID: 8824222

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

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

Vicente98: Vicente M, Gomez MJ, Ayala JA (1998). "Regulation of transcription of cell division genes in the Escherichia coli dcw cluster." Cell Mol Life Sci 54(4);317-24. PMID: 9614967

Wang01e: Wang W, Maniar M, Jain R, Jacobs J, Trias J, Yuan Z (2001). "A fluorescence-based homogeneous assay for measuring activity of UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase." Anal Biochem 290(2);338-46. PMID: 11237337

Westphal12: Westphal K, Langklotz S, Thomanek N, Narberhaus F (2012). "A trapping approach reveals novel substrates and physiological functions of the essential protease FtsH in Escherichia coli." J Biol Chem 287(51);42962-71. PMID: 23091052

Young91: Young K, Silver LL (1991). "Leakage of periplasmic enzymes from envA1 strains of Escherichia coli." J Bacteriol 173(12);3609-14. PMID: 1904854

Young95: Young K, Silver LL, Bramhill D, Cameron P, Eveland SS, Raetz CR, Hyland SA, Anderson MS (1995). "The envA permeability/cell division gene of Escherichia coli encodes the second enzyme of lipid A biosynthesis. UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase." J Biol Chem 270(51);30384-91. PMID: 8530464

Zeng13: Zeng D, Zhao J, Chung HS, Guan Z, Raetz CR, Zhou P (2013). "Mutants resistant to LpxC inhibitors by rebalancing cellular homeostasis." J Biol Chem 288(8);5475-86. PMID: 23316051

Other References Related to Gene Regulation

Eraso14: Eraso JM, Markillie LM, Mitchell HD, Taylor RC, Orr G, Margolin W (2014). "The Highly Conserved MraZ Protein Is a Transcriptional Regulator in Escherichia coli." J Bacteriol 196(11);2053-66. PMID: 24659771

Gohler11: Gohler AK, Kokpinar O, Schmidt-Heck W, Geffers R, Guthke R, Rinas U, Schuster S, Jahreis K, Kaleta C (2011). "More than just a metabolic regulator - elucidation and validation of new targets of PdhR in Escherichia coli." BMC Syst Biol 5(1);197. PMID: 22168595

Ishino89: Ishino F, Jung HK, Ikeda M, Doi M, Wachi M, Matsuhashi M (1989). "New mutations fts-36, lts-33, and ftsW clustered in the mra region of the Escherichia coli chromosome induce thermosensitive cell growth and division." J Bacteriol 171(10);5523-30. PMID: 2676977

MendozaVargas09: Mendoza-Vargas A, Olvera L, Olvera M, Grande R, Vega-Alvarado L, Taboada B, Jimenez-Jacinto V, Salgado H, Juarez K, Contreras-Moreira B, Huerta AM, Collado-Vides J, Morett E (2009). "Genome-wide identification of transcription start sites, promoters and transcription factor binding sites in E. coli." PLoS One 4(10);e7526. PMID: 19838305

Mitchell03: Mitchell JE, Zheng D, Busby SJ, Minchin SD (2003). "Identification and analysis of 'extended -10' promoters in Escherichia coli." Nucleic Acids Res 31(16);4689-95. PMID: 12907708


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 18.5 on Sat Nov 22, 2014, BIOCYC14B.