Genetic and Biochemical Analyses ofEscherichia coliMutants Altered in the Temperature-Dependent Regulation of Membrane Lipid Composition

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Abstract
Two mutants of E. coli altered in the regulation of membrane lipid composition by temperature were previously studied. One class (represented by the fabF1 allele) fails to regulate upon temperature shift and is defective in cis-vaccenic acid synthesis owing to the lack of the fatty acid elongation enzyme .beta.-ketoacyl-acyl carrier protein synthase II (EC 2.3.1.41). A second class of mutant, given the phenotypic designation Vtr, overproduces cis-vaccenic acid at all temperatures and hence is altered in temperature regulation. The Vtr and fabF1 mutations show very tight genetic linkage. The Vtr lesion is allelic to the fabF1 mutation since the presence of the fabF1 mutation in merodiploid strains carrying the Vtr or fabF+ alleles results in fatty acid compositions intermediate between those of the 2 monoploid strains. Merodiploids carrying the fabF+ and Vtr alleles likewise show an intermediate composition. These results indicate intra-allelic complementation. The 2 recently discovered E. coli proteins that form mixed disulfide cross-links to acyl carrier protein are directly demonstrated to be .beta.-ketoacyl-acyl carrier protein synthases I and II. The fabF1 strains produce a synthase II band of altered electrophoretic mobility, indicating that the fabF locus is the structural gene for synthase II. The synthase II of Vtr strains is abnormally sensitive to cerulenin, an antibiotic that specifically inhibits synthase I and II. This increased sensitivity is readily demonstrated in vivo, but in vitro it was impossible to detect an increased sensitivity of the Vtr synthase II to cerulenin nor was any other kinetic or structural alteration in the enzyme detected. These results were interpreted in terms of specific interactions of synthase II with other cellular components which occur in vivo but are not duplicated in vitro.