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Characterization of E. coli mutants resistant to LpxC inhibitors

The emergence of multidrug resistant Gram‐negative bacteria requires the search for antibiotics with novel targets. One such target is lipid A, the anchor of lipopolysaccharide on the surface of Gram‐negative bacteria. Several potent and bactericidal inhibitors have been developed recently targeting LpxC, the deacetylase that catalyzes the second and committed step of lipid A biosynthesis. Here, we study mechanisms of resistance to these inhibitors by generating spontaneously resistant mutants that are ~200 fold resistant to all known LpxC inhibitors. These resistant mutants are isolated at a low rate and in two‐stages. All resistant mutants have two chromosomal point mutations that account for all resistance additively: one in thrS, the Thr‐tRNA ligase and the other in fabZ, a dehydrase in fatty acid biosynthesis that shares a common substrate with LpxA, the enzyme preceding LpxC in lipid A biosynthesis. The point mutations in thrS and fabZ decrease enzymatic activity. We speculate the mutation in thrS decreases protein synthesis and therefore slows cell growth, allowing the toleration of decreased lipid A biosynthesis, and the mutation in fabZ increases the amount of LpxC substrate that could compete with inhibitors. Understanding these resistance mechanisms has important implications in drug development and membrane homeostasis. This research was funded by N.I.H. grant GM51310 to C.R.H. Raetz.