Outer membrane permeability barrier in Escherichia coli mutants that are defective in the late acyltransferases of lipid A biosynthesis

Abstract

The tight packing of six fatty acids in the lipid A constituent of lipopolysaccharide (LPS) has been proposed to contribute to the unusually low permeability of the outer membrane of gram-negative enteric bacteria to hydrophobic antibiotics. Here it is shown that the Escherichia coli msbB mutant, which elaborates defective, penta-acylated lipid A, is practically as resistant to a representative set of hydrophobic solutes (rifampin, fusidic acid, erythromycin, clindamycin, and azithromycin) as the parent-type control strain. The susceptibility index, i.e., the approximate ratio between the MIC for the msbB mutant and that for the parent-type control, was maximally 2.7-fold. In comparison, the rfa mutant defective in the deep core oligosaccharide part of LPS displayed indices ranging from 20 to 64. The lpxA and lpxD lipid A mutants had indices higher than 512. Furthermore, the msbB mutant was resistant to glycopeptides (vancomycin, teicoplanin), whereas the rfa, lpxA, and lpxD mutants were susceptible. The msbB htrB double mutant, which elaborates even-more-defective, partially tetra-acylated lipid A, was still less susceptible than the rfa mutant. These findings indicate that hexa-acylated lipid A is not a prerequisite for the normal function of the outer membrane permeability barrier.

FIG. 1

Structures of the lipid A part of the LPS of wild-type E. coli (A) and the msbB mutant of E. coli, which lacks the myristic acid (C14) residue (B). The msbB htrB double mutant also lacks part of the lauric acid residue (C12) (see text). Acylations catalyzed by MsbB and HtrB are shown in panel A.

FIG. 2

MICs of rifampin (A), fusidic acid (B), erythromycin (C), clindamycin (D), azithromycin (E), and vancomycin (F) for E. coli mutants defective in the synthesis of LPS core oligosaccharide or lipid A as well as for their parent-type controls. Susceptibility determinations were performed at 28°C (○) and 37°C (■). Lanes: 1, strain IH3080 (smooth, encapsulated control); 2, strain SM105 (K-12 wild-type control); 3, strain MLK1067 (msbB); 4, strain MLK53 (htrB); 5, strain MLK986 (msbB htrB); 6, strain D21f2 (rfa, LPS chemotype Re); 7, strain CDH23-213 (lpxD); and 8, strain SM101 (lpxA). Susceptibility of a gram-positive bacterium, M. luteus ATCC 9341 (lanes 9), is shown for comparison.