The effect of epetraborole on the transcriptome and proteome profiles of an Escherichia coli strain overexpressing leuS, Leucyl-tRNA Synthetase

Abstract

Epetraborole (EP) is a boron-containing antibiotic known for its effectiveness against gram-negative enteric bacteria and Mycobacterium species. It is designed to bind and inhibit the LeuS enzyme (Leucyl-tRNA Synthetase), which is encoded by the essential gene leuS in Escherichia coli. EP inhibits protein translation, impeding bacterial growth. However, when leuS is overexpressed in a recombinant plasmid, the amount of EP required for growth inhibition needs to be increased. This study explored the impact of EP on the transcriptome and proteome of E. coli overexpressing leuS, aiming to reveal additional gene and pathway insights beyond LeuS, shedding light on the biochemical players orchestrating the bacterium’s molecular response. 2D-PAGE Proteomics analysis identified four differentially regulated proteins influenced by EP in the leuS overexpression strain. Notably, LeuA and DeoA emerged as identified proteins. EP may affect LeuA in the cells overexpressing LeuS, which could result in truncated LeuA protein variants. Transcriptomics analyses, based on microarray data, revealed 23 up-regulated and 9 down-regulated genes responding to EP in the overexpression strain (p2). Based on the statistical analyses, the first five up-and down-regulated genes showing the highest fold differences in their mRNA levels are yiaW, mglB, narH, ybiO, flgB and yhdY, deoR, recX, yobB, potF, respectively. Analyses using the Omics Dashboard pathway and String indicate that the EP effect on the leuS overexpressing strain mainly induces alterations in the expression of genes related to the cell exterior, regulation, and response to stimuli. It is suggested that EP and higher levels of LeuS may interfere with the translational and transcriptional regulation of the expression of the leuA gene, which encodes the first enzyme, 2-isopropylmalate synthase, in L-leucine biosynthesis. This study offers new insights into the effects of EP on the bacterium, specifically when the level of the aminoacyl-tRNA synthetase LeuS is increased.