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Isolation and Chemical Characterization of Lipid A from Gram-negative Bacteria
Published on: September 16, 2013
Regulated covalent modifications of lipid A
1Department of Biochemistry, Duke University Medical Center, Durham, NC, USA. raetz@biochem.duke.edu
Journal of Endotoxin Research
|August 25, 2001
Summary
Pathogenic bacteria modify lipid A for virulence. Researchers identified a novel enzyme, LpxO, responsible for 2-hydroxymyristate modification of lipid A, crucial for bacterial survival and potential therapeutic targeting.
Area of Science:
- Microbiology
- Biochemistry
- Molecular Biology
Background:
- Regulated covalent modifications of lipid A are crucial for the virulence of Gram-negative bacteria.
- Genes like pagP and pagL encode outer membrane enzymes involved in lipid A modification, impacting resistance to antimicrobial peptides.
- Salmonella typhimurium synthesizes S-2-hydroxymyristate modified lipid A in a PhoP/PhoQ-dependent manner.
Purpose of the Study:
- To identify the enzyme responsible for the 2-hydroxylation of lipid A in Salmonella.
- To characterize the function and localization of the identified enzyme.
Main Methods:
- Bioinformatic searches using dioxygenase sequences to identify homologous genes.
- Cloning and expression of the S. typhimurium aspartyl beta-hydroxylase homologue (lpxO) in Escherichia coli K-12.
- Analysis of lipid A modifications in E. coli expressing lpxO using mass spectrometry.
Main Results:
- A novel open reading frame, lpxO, with similarity to mammalian aspartyl beta-hydroxylases was identified in bacteria producing 2-hydroxyacylated lipid A.
- Expression of lpxO in E. coli induced oxygen-dependent formation of 2-hydroxymyristate-modified lipid A.
- LpxO is proposed to be an inner membrane enzyme catalyzing Fe(2+)/ascorbate/alpha-ketoglutarate dependent hydroxylation of lipid A.
Conclusions:
- LpxO is the enzyme responsible for 2-hydroxymyristate modification of lipid A in S. typhimurium.
- This modification is essential for bacterial virulence and survival.
- 2-hydroxymyristoyl coenzyme A, derived from released 2-hydroxymyristate, may inhibit host protein N-myristoyl transferase, suggesting a novel virulence mechanism.
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