Cell wall stress response proteins in B. subtilis and their potential as new antibiotic targets: YtrB and YtrE, subunits of a putative ABC transporter

Sammanfattning

Multi-resistant bacteria exist worldwide and are a serious problem with 25 000 deaths per year only in the EU. The effort of finding new antibiotics has also reduced over the past decades due to resistance being established even before the drug reaches the market. A relatively new idea is to design antibiotics with multiple targets and/or multiple effects. One of these targets with multiple effects is the cell wall stress response. Previous investigations used proteomic studies of the cell wall stress response in Bacillus subtilis (B. subtilis) and found different marker proteins for different classes of antibiotics. In this study, we focused on the cell wall synthesis marker proteins YtrB and YtrE, which are annotated as nucleotide-binding subunits of a putative ABC transporter. The aim was to study their localisation, importance for growth at different temperatures, and their impact on antibiotic sensitivity to evaluate the transporter’s potential as a novel drug target. GFP fusions and deletion mutants were constructed and used in growth curve experiments, bacterial cytological profiling (BCP), and minimal inhibitory concentration (MIC) assays. Neither YtrB nor YtrE were of high importance for the growth in B. subtilis at different temperatures. They were, however, important for the resistance against the lantibiotic nisin and the beta lactams ampicillin and ertapenem. By connecting the localisation of YtrB in both rigid and fluid regions of the membrane to previous studies and to our MIC results, it can be hypothesised that the ABC transporter may have multiple functions. Among these are the function as an importer of precursor molecules for cell wall synthesis and/or the function as a membrane-bound remover or inhibitor of beta lactams. The ABC transporter has, as such, the potential of being a target for new potentiators: sensitizers to decrease the cell wall thickness or increase the activity of beta lactams. However, more studies need to be done with additional antibiotics and strains (a reporter fusion of the promoter of the ytr operon, whole ytr operon deletion, and N-terminal GFP fusion to YtrE) to determine their potential use within healthcare.