Studies on the Uridine Diphosphate-Galactose:Lipopolysaccharide Galactosyltransferase Reaction Using a Fatty Acid Mutant of Escherichia coli

Abstract

Abstract The UDP-galactose:lipopolysaccharide galactosyltransferase reaction has been studied using an unsaturated fatty acid auxotroph of Escherichia coli. The unsaturated fatty acid residues of the E. coli membrane phospholipid have been varied with respect to chain length, position, number, and configuration (cis versus trans) of the ethylenic bond. Although the galactose content of the lipopolysaccharide from cells with membrane phospholipid altered in this way is not appreciably reduced, the reaction itself is found to be sensitive to changes in the fatty acid structure of the phospholipid. The reaction was measured in EDTA-treated intact cells, or using supernatant enzyme and cell envelopes, or membranes, as a source of galactose-deficient acceptor. In general, the activity was greater under conditions known to restrict the packing of the fatty acid residues of phospholipid; for example, shorter chain length, an ethylenic bond near the carboxyl terminus, and cis- rather than trans-unsaturated fatty acids. The activity was also reduced to a much greater extent at lower assay temperatures (l37°) using preparations containing trans-unsaturated fatty acid than with preparations containing the corresponding cis-unsaturated fatty acid. Arrhenius plots of the activity using cell envelopes show a discontinuity when the unsaturated fatty acid is cis-Δ9-16:1, cis-Δ9-18:1, or cis-Δ11-18:1, but not when the corresponding trans-unsaturated fatty acid is used. The temperature at which the discontinuity occurs is 18–23° for all three cis-unsaturated fatty acids.