Acyl-[acyl-carrier-protein]-UDP-N-acetylglucosamine O-acyltransferase

 

UDP-N-acetylglucosamine (UDP-GlcNAc) acyltransferase catalyzes the first step in the biosynthesis of lipid A, the hydrophobic anchor of lipopolysaccharide in Gram-negative bacteria. This enzyme, the product of the lpxA gene, transfers an R-3-hydroxyacyl chain fromR-3-hydroxyacyl-acyl carrier protein (ACP) to the glucosamine 3-OH of UDP-GlcNAc. The acylation of UDP-GlcNAc is characterized by an unfavorable equilibrium constant (0.01). Therefore, the second reaction of lipid A biosynthesis, in which the LpxA product UDP-3-O-(R-3-hydroxyacyl)-GlcNAc is deacetylated, is the first irreversible step of the pathway. Lipid A is required for growth of E. coli and most other Gram-negative bacteria. Lipid A is also necessary for maintaining the integrity of the outer membrane as a barrier to toxic chemicals. Furthermore, lipid A is a potent activator of innate immunity in animal systems. The study of the enzymes involved in lipid A biosynthesis should therefore prove useful for the development of new anti-infective drugs.

 

Reference Protein and Structure

Sequence
P0A722 UniProt (2.3.1.129) IPR010137 (Sequence Homologues) (PDB Homologues)
Biological species
Escherichia coli K-12 (Bacteria) Uniprot
PDB
1lxa - UDP N-ACETYLGLUCOSAMINE ACYLTRANSFERASE (2.6 Å) PDBe PDBsum 1lxa
Catalytic CATH Domains
2.160.10.10 CATHdb (see all for 1lxa)
Click To Show Structure

Enzyme Reaction (EC:2.3.1.129)

O-[S-(3R)-hydroxytetradecanoylpantetheine-4-phosphoryl]serine(1-) residue
CHEBI:78474ChEBI
+
UDP-N-acetyl-alpha-D-glucosamine(2-)
CHEBI:57705ChEBI
→
O-(pantetheine-4'-phosphoryl)serine(1-) residue
CHEBI:64479ChEBI
+
UDP-3-O-[(3R)-3-hydroxytetradecanoyl]-N-acetylglucosamine(2-)
CHEBI:61494ChEBI
Alternative enzyme names: UDP-N-acetylglucosamine acyltransferase, Uridine diphosphoacetylglucosamine acyltransferase, Acyl-[acyl-carrier-protein]-UDP-N-acetylglucosamine O-acyltransferase, (R)-3-hydroxytetradecanoyl-[acyl-carrier-protein]:UDP-N-acetylglucosamine 3-O-(3-hydroxytetradecanoyl)transferase, (R)-3-hydroxytetradecanoyl-[acyl-carrier-protein]:UDP-N-acetyl-alpha-D-glucosamine 3-O-(3-hydroxytetradecanoyl)transferase,

Enzyme Mechanism

Introduction

His125 deprotonates UDP-N-acetyl-D-glucosamine hydroxyl group, initiating a nucleophilic attack on the carbonyl carbon of the ACP in an addition reaction. The oxyanion collapses, eliminating ACP with concomitant deprotonation of His125.

Catalytic Residues Roles

UniProt PDB* (1lxa)
Gly143 (main-N) Gly143A (main-N) Backbone amide forms oxyanion hole to stabilise build up of negative charge during the reaction. hydrogen bond donor, electrostatic stabiliser
Asp126 Asp126A Putatively aids His125 in its function as a general base. hydrogen bond acceptor, hydrogen bond donor, electrostatic stabiliser
His125 His125A Acts as a general base to deprotonate the substrate hydroxyl group. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

proton transfer, bimolecular nucleophilic addition, overall reactant used, intermediate formation, unimolecular elimination by the conjugate base, overall product formed, native state of enzyme regenerated, intermediate terminated

References

  1. Wyckoff TJ et al. (1999), J Biol Chem, 274, 27047-27055. The Active Site of Escherichia coliUDP-N-acetylglucosamine Acyltransferase: CHEMICAL MODIFICATION AND SITE-DIRECTED MUTAGENESIS. DOI:10.1074/jbc.274.38.27047. PMID:10480918.
  2. Williams AH et al. (2007), Proc Natl Acad Sci U S A, 104, 13543-13550. Structural basis for the acyl chain selectivity and mechanism of UDP-N-acetylglucosamine acyltransferase. DOI:10.1073/pnas.0705833104. PMID:17698807.
  3. Ulaganathan V et al. (2007), J Mol Biol, 369, 305-312. Nucleotide Substrate Recognition by UDP-N-acetylglucosamine Acyltransferase (LpxA) in the First Step of Lipid A Biosynthesis. DOI:10.1016/j.jmb.2007.03.039. PMID:17434525.
  4. Williams AH et al. (2006), Proc Natl Acad Sci U S A, 103, 10877-10882. Structure of UDP-N-acetylglucosamine acyltransferase with a bound antibacterial pentadecapeptide. DOI:10.1073/pnas.0604465103. PMID:16835299.
  5. Sweet CR et al. (2004), J Biol Chem, 279, 25411-25419. Enzymatic Synthesis of Lipid A Molecules with Four Amide-linked Acyl Chains: LpxA ACYLTRANSFERASES SELECTIVE FOR AN ANALOG OF UDP-N-ACETYLGLUCOSAMINE IN WHICH AN AMINE REPLACES THE 3''-HYDROXYL GROUP. DOI:10.1074/jbc.m400597200. PMID:15044493.
  6. Jain NU et al. (2004), J Mol Biol, 343, 1379-1389. Rapid Analysis of Large Protein–Protein Complexes Using NMR-derived Orientational Constraints: The 95kDa Complex of LpxA with Acyl Carrier Protein. DOI:10.1016/j.jmb.2004.08.103. PMID:15491619.
  7. Lee BI et al. (2003), Proteins, 53, 772-774. Crystal structure of UDP-N-acetylglucosamine acyltransferase fromHelicobacter pylori. DOI:10.1002/prot.10436. PMID:14579368.

Step 1. His125 deprotonates UDP-N-acetyl-D-glucosamine hydroxyl group, initiating a nucleophilic attack on the carbonyl carbon of the ACP in an addition reaction.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His125A hydrogen bond acceptor, hydrogen bond donor
Asp126A hydrogen bond acceptor
Gly143A (main-N) hydrogen bond donor, electrostatic stabiliser
His125A proton acceptor

Chemical Components

proton transfer, ingold: bimolecular nucleophilic addition, overall reactant used, intermediate formation

Step 2. The oxyanion collapses, eliminating ACP with concomitant deprotonation of His125.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His125A hydrogen bond donor
Asp126A hydrogen bond donor, electrostatic stabiliser
Gly143A (main-N) hydrogen bond donor, electrostatic stabiliser
His125A proton donor

Chemical Components

proton transfer, ingold: unimolecular elimination by the conjugate base, overall product formed, native state of enzyme regenerated, intermediate terminated
Download: Image, Marvin File

Step 1. His125 deprotonates UDP-N-acetyl-D-glucosamine hydroxyl group, initiating a nucleophilic attack on the carbonyl carbon of the ACP in an addition reaction.

Step 2. The oxyanion collapses, eliminating ACP with concomitant deprotonation of His125.

Products.

Contributors

Gemma L. Holliday, Daniel E. Almonacid, Gail J. Bartlett, Anna Waters, Craig Porter