PMID:8530464

Young, K, Silver, LL, Bramhill, D, Cameron, P, Eveland, SS, Raetz, CR, Hyland, SA and 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. 270:30384-91

Abstract

The envA gene of Escherichia coli has been shown previously to be essential for cell viability (Beall, B. and Lutkenhaus, J. (1987) J. Bacteriol. 169, 5408-5415), yet it encodes a protein of unknown function. Extracts of strains harboring the mutant envA1 allele display 3.5-18-fold reductions in UDP-3-O-acyl-N-acetylglucosamine deacetylase specific activity. The deacetylase is the second enzymatic step of lipid A biosynthesis. The structural gene coding for the deacetylase has not been assigned. In order to determine if the envA gene encodes the deacetylase, envA was cloned into an isopropyl-1-thio-beta-D-galactopyranoside-inducible T7-based expression system. Upon induction, a protein of the size of envA was highly overproduced, as judged by SDS-PAGE. Direct deacetylase assays of cell lysates revealed a concomitant approximately 5,000-fold overproduction of activity. Assays of the purified, overproduced EnvA protein demonstrated a further approximately 5-fold increase in specific activity. N-terminal amino acid sequencing of the purified protein showed that the first 20 amino acids matched the predicted envA nucleotide sequence. Contaminating species were present at less than 1% of the level of the EnvA protein. Thus, envA is the structural gene for UDP-3-O-acyl-GlcNAc deacetylase. Based on its function in lipid A biosynthesis, we propose the new designation lpxC for this gene.

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Annotations

Gene product	Qualifier	GO ID	GO term name	Evidence Code	with/from	Aspect	Notes	Status
LpxC		GO:0008759	UDP-3-O-(3-hydroxymyristoyl) N-acetylglucosamine deacetylase activity	IDA: Inferred from Direct Assay		F	Assayed using [alpha-32P]UDP-3-O-(R-3-hydroxymyristoyl)-GlcNAc as the substrate. Product was separated from the reactant by thin layer chromatography and quantified by scintillation counting. Substrate is not commercially available but its preparation is described.	complete
LpxC		GO:0009245	lipid A biosynthetic process	IMP: Inferred from Mutant Phenotype		P	Extracts prepared from strains with the lpxC101 (envA1) mutation have greatly reduced levels of UDP-3-O-acyl-N-acetylglucosamine deacetylase activity (Tables II and III). This activity is greatly enhanced by overexpression of the lpxC gene from a T7 promoter (Table III).	complete
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