title: Isolation and structural analysis of two lipid A precursors from a KDO deficient mutant of Salmonella typhimurium differing in their hexadecanoic acid content.
abstract: The extraction, purification and structural characterization of two lipid A precursors (Ia and Ib) differing only in one hexadecanoic acid are described. Both precursors were synthesized at elevated temperatures by a new mutant of Salmonella typhimurium (mutant Ts5) which is conditionally defective in synthesis of the 3-deoxy-D-manno-octulosonic acid region of lipopolysaccharides. Both precursors were purified by repeated phenol/chloroform/petroleum ether (PCP) extractions followed by thin layer chromatography. The precursor preparation was free of lipopolysaccharides and phospholipids and contained less than 0.1% protein. Structural analysis which included chemical degradation procedures as well as positive ion laser desorption (LDMS) mass spectroscopy of dephosphorylated lipid A precursors showed together that precursor Ia represents a diphosphorylated glucosamine disaccharide containing two ester, two amide-linked residues of 3-hydroxytetradecanoic acid and lacks the ester-linked dodecanoic, tetradecanoic and hexadecanoic acid as well as 3-deoxy-D-manno-octulosonic acid. Precursor Ib has the same basic structure as precursor Ia, but contains in addition one mol of hexadecanoic acid per mol disaccharide which is linked to the 3-hydroxy group of the amide-bound 3-hydroxy-tetradecanoic acid of the reducing, terminal glucosamine residue. The structure of precursor Ib supports the conclusion that hexadecanoic acid incorporation occurs at an early stage in lipid A biosynthesis prior to the attachment of 3-deoxy-D-manno-octulosonic acid and/or other polar substituents.
mesh_terms: D002848:Chromatography, DEAE-Cellulose; D002854:Chromatography, Paper; D002855:Chromatography, Thin Layer; D006017:Glycolipids; D008050:Lipid A / Q000031:analogs & derivatives* / Q000032:analysis / Q000302:isolation & purification; D013058:Mass Spectrometry; D009154:Mutation; D010169:Palmitic Acids / Q000032:analysis*; D012486:Salmonella typhimurium / Q000032:analysis* / Q000235:genetics; D013400:Sugar Acids / Q000032:analysis*
chemical_list: D006017:Glycolipids; D008050:Lipid A; D010169:Palmitic Acids; D013400:Sugar Acids; C040142:lipid A precursors, bacterial; C002532:2-keto-3-deoxyoctonate