Isolation of Mutants Conditionally Blocked in the Biosynthesis of the 3‐Deoxy‐d‐manno‐octulosonic‐acid—Lipid‐A Part of Lipopolysaccharides Derived from Salmonella typhimurium

Abstract

A procedure is described for the selection of conditional 3-deoxy-D-manno-octulosonic-acid-lipid A mutants which depends on temperature sensitivity for the synthesis of complete lipopolysaccharide and for growth. Using this procedure new types of mutants were isolated which cease growth and accumulate lipid A precursors following a shift to nonpermissive temperatures. All precursor molecules differ in their charge as judged by DEAE-cellulose chromatography. While they all contain glucosamine, phosphate and 3-hydroxymyristic acid, they lack detectable 3-deoxy-D-manno-octulosonic acid (dOclA) and the nonhydroxylated fatty acids of the complete lipid A structure. Three mutants were conditionally defective in dOclA metabolism, but one seems to be blocked at a relatively early step in lipid A synthesis. The phenotypes of all these mutants appear to be due to single mutations by reversion analysis and by characterization of the temperature-resistant revertants. Studies of these mutants may shed light on the essential role of the complete dOclA-lipid A part of lipopolysaccharides in membrane function.