An integrated quantitative proteomic and metabolomics approach to reveal the negative regulation mechanism of LamB in antibiotics resistance

Abstract

Previously, a maltose-specific channel porin, LamB was found to be associate with multi-drug resistance in a lamB deleted strain, but the exact mechanisms require further elucidation. Herein, differential protein expression between the Escherichia coli mutant strain ΔlamB and the wild type strain BW25113, with and without ciprofloxacin (CFLX), was identified using iTRAQ based liquid chromatography-tandem mass spectrometry (LC-MS/MS); while differential metabolite expression was examined using gas chromatography–mass spectrometry (GC–MS). Further Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that lamB deletion leads to a decrease in several key metabolic pathways such as tricarboxylic acid (TCA) cycle, pentose phosphate pathway and glycolysis/gluconeogenesis. When examining the ΔlamB strain without CFLX, many aminoacyl-tRNA biosynthesis and pyrimidine metabolism-related proteins were unaltered, but the addition of CFLX resulted in reduced levels. These findings indicate that a lamB deletion may confer antibiotic resistance by relieving the pressure of protein translation and DNA replication. To further examine antibiotic resistance, exogenous metabolites, including maltose, and several amino acids metabolites were evaluated to determine whether the resistance level could be reduced in the presence of CFLX. The obtained results indicate that lamB knockout may increase bacterial antibiotics resistance by decreasing metabolic pathway activity levels. Biological significanceAn integrated metabonomic-proteomic method was performed to systematically compare the profiles of metabolites and proteins between ΔlamB and its wild type strain, with and without ciprofloxacin (CFLX) treatment. Following bioinformatics analysis showed that lamB deletion in CFLX stress leads to the decreasing of several key metabolic pathways. Many amino acid-tRNA biosynthesis and pyrimidine metabolism related proteins didn't change in ΔlamB strain but largely decreased after treated with CFLX. Further exogenous metabolites addition assay reveals that maltose and several amino acids metabolites contribute to the CFLX resistance mediated by LamB. Our results indicate that the down-regulation of LamB may increase bacterial antibiotics resistance by decreasing the intracellular metabolism pathways.