DEFECTIVE BIOSYNTHESIS OF THE LIPID-A COMPONENT OF TEMPERATURE-SENSITIVE FIRA (OMSA) MUTANT OF ESCHERICHIA-COLI

The biosynthesis of lipid A component was shown to be defective in a temperature-sensitive firA mutant of Escherichia coli. Cells were biosynthetically labelled with [C-14]acetate and incorporation of radioactivity into the glycerophospholipid compared to lipid A fractions was measured. The lipid A/glycerophospholipid biosynthesis ratio of the firA mutant at 37-degrees-C was approximately 50%, and at the nonpermissive temperature of 42-degrees-C was less than 20% of that observed in the corresponding wild-type strain. Analysis of radiolabelled lipid A 4'-monophosphate derivatives and glycerophospholipids by thin-layer chromatography revealed that the firA mutant at 42-degrees-C elaborated an altered lipid A, and its phosphatidylglycerol content was low. The chemical composition of the extracted lipopolysaccharides differed significantly between the firA and the wild-type strain only in the proportion of hexadecanoic acid, which was minimal in the wild type grown at 37-degrees-C and 42-degrees-C and in firA lipopolysaccharide grown at 37-degrees-C. In the firA mutant lipopolysaccharide produced at 42-degrees-C, hexadecanoic acid was present in approximately every third molecule, attached to the hydroxyl group of the amide-linked (R)-3-hydroxytetradecanoic acid at the reducing glucosamine of lipid A. Inspection of dephosphorylated free lipid A preparations by laser-desorption mass spectrometry confirmed that significant amounts of heptaacyl lipid A was elaborated by the firA strain grown at 42-degrees-C.