Overexpression of lpxT Gene in Escherichia coli Inhibits Cell Division and Causes Envelope Defects without Changing the Overall Phosphorylation Level of Lipid A

Abstract

LpxT is an inner membrane protein that transfers a phosphate group from the essential lipid undecaprenyl pyrophosphate (C-55PP) to the lipid A moiety of lipopolysaccharide, generating a lipid A tris-phosphorylated species. The protein is encoded by the non-essential lpxT gene, which is conserved in distantly related Gram-negative bacteria. In this work, we investigated the phenotypic effect of lpxT ectopic expression from a plasmid in Escherichia coli. We found that lpxT induction inhibited cell division and led to the formation of elongated cells, mostly with absent or altered septa. Moreover, the cells became sensitive to detergents and to hypo-osmotic shock, indicating that they had cell envelope defects. These effects were not due to lipid A hyperphosphorylation or C-55PP sequestering, but most likely to defective lipopolysaccharide transport. Indeed, lpxT overexpression in mutants lacking the L,D-transpeptidase LdtD and LdtE, which protect cells with outer membrane defects from osmotic lysis, caused cell envelope defects. Moreover, we found that pyrophosphorylated lipid A was also produced in a lpxT deletion mutant, indicating that LpxT is not the only protein able to perform such lipid A modification in E. coli.

Flux redistribution of central carbon metabolism for efficient production of l‐tryptophan in Escherichia coli

“…A possible explanation is that the Glf enzyme is a glucose transporter with high uptake efficiency, and too strong an expression of Glf may increase the intracellular glucose concentration, thus interfering with normal metabolism (Lara et al, 2008) and cell growth (Kwiatkowska et al, 2008). In addition, overexpression of membrane protein is likely to cause toxicity (Gubellini et al, 2011) and cell envelope defects (Falchi et al, 2020). As the introduction of strongly expressed glf gene, including P M1‐37 ‐ glf and P M1‐93 ‐ glf , resulted in a significantly lower glucose consumption and biomass than those of introduction of P M1‐12 ‐ glf , we selected the strain with P M1‐12 ‐ glf for further genetic modification.…”