An inner membrane dioxygenase that generates the 2-hydroxymyristate moiety of Salmonella lipid A.

摘要: The lipid A residues of certain Gram-negative bacteria, including most strains Salmonella and Pseudomonas, are esterified with one or two secondary S-2-hydroxyacyl chains. S-2 hydroxylation process is O2-dependent in vivo, but the relevant enzymatic pathways have not been fully characterized because vitro assays developed. We previously reported that expression lpxO gene confers upon Escherichia coli K-12 ability to synthesize 2-hydroxymyristate modified (Gibbons, H. S., Lin, Cotter, R. J., Raetz, C. J. Biol. Chem. 275, 32940–49, 2000). now demonstrate inactivation lpxO, which encodes a putative Fe2+/O2/α-ketoglutarate-dependent dioxygenase, abolishes S-2-hydroxymyristate formation S. typhimurium. Membranes E. expressing able hydroxylate Kdo2-[4′-32P]-lipid presence Fe2+, O2, α-ketoglutarate, ascorbate Triton X-100. Fe2+ chelator 2,2′-bipyridyl inhibits reaction. product generated mono-hydroxylated Kdo2-lipid derivative. [4′-32P]-lipid released by mild acid hydrolysis from migrates authentic S-2-hydroxlyated isolated 32P-labeled typhimurium cells. Electrospray ionization mass spectrometry gas chromatography/mass consistent 2-hydroxylation 3′-secondary myristoyl chain A. LpxO contains predicted trans-membrane helices (one at each end protein), its active site likely faces cytoplasm. an unusual example integral membrane protein member dioxygenase family.