INVOLVEMENT OF WAAY, WAAQ, AND WAAP IN THE MODIFICATION OF ESCHERICHIA-COLI LIPOPOLYSACCHARIDE AND THEIR ROLE IN THE FORMATION OF A STABLE OUTER-MEMBRANE

The waaY, waaQ, and waaP genes are located in the central operon of th e waa (formerly rfa.) locus on the chromosome of Escherichia coli. Thi s locus contains genes whose products are involved in the assembly of the core region of the lipopolysaccharide molecule. In the R1 core pro totype strain, E. coli F470, there are nine genes in this operon, and all but waaY, waaQ, and waaP have been assigned function, In this stud y, the waaY, waaQ, and waaP genes were independently mutated by insert ion of a non-polar antibiotic resistance cassette, and the structures of the resulting mutant core oligosaccharides were determined by chemi cal analyses and phosphorus-nuclear magnetic resonance spectroscopy. A ll three of these mutations were shown to affect the modification of t he heptose region of the core, a region whose structure is critical to outer membrane stability. Mutation of waaY resulted in a core oligosa ccharide devoid of phosphate on HepII. Mutation of waaQ resulted in lo ss of the branch HepIII residue on HepII and impeded the activity of W aaY. Mutation of waaP resulted in loss of phosphoryl substituents on H epI and obviated WaaQ and WaaY activity. Only mutation of waaP resulte d in hypersensitivity to novobiocin and sodium dodecyl sulfate, a char acteristic of deep-rough mutations.