Proteomics Study On The Mechanism Of Drug Resistance In Bacteria

Bacterial drug resistance seriously threatens public health.With the emergence of superbugs and resistant plasmids,such as MRSA,NDM-1,MCR-1,the study of the bacterial resistance mechanism is urgently needed.Currently,it has made certain progress in the studies of bacterial drug resistance,whereas most of the relevant studies focused on the isolation and identification of clinical multi-drugresistant bacteria.Although the single target-based methods can be used to reveal a few mechanisms,it is impossible to understand fully the correlation between various bacterial resistance mechanisms considering the complex genetic background of clinical drug-resistant bacteria.Thus,in this study,we domesticated bacteria(Escherichia coli)with antibiotics(ciprofloxacin and ampicillin)in the laboratory,and selected the low,medium,and high drugresistant bacteria as a dynamic model.Then,quantitative proteomic technology combined with functional validations was performed to uncover the evolutionary mechanism of antibiotic resistance in bacteria.The data showed that bacteria systematically regulate the expression levels of proteins to change the conversion direction of material and energy,and thus enhance bacterial resistance to antibiotics.The specific mechanisms are as follows: enhancement of energy synthesis and accumulation by maintaining the redox balance,improvement of antioxidant activity,increase of the antagonism in antibiotic targets,enhancement of biofilms formation by reducing cell motility and raising quorum sensing,improvement of drug efflux and decrease of antibiotics permeation.Moreover,we also found that the evolution process of bacterial resistance has the characteristics of continuity in whole and stage-by-stage in part.In addition to the common continuous evolution mechanism,we also found that there were differences in the energy synthesis modes and levels in different resistance stages,suggesting the stage equilibrium mechanism in the evolution process.Moreover,there are differences in antioxidant patterns among different types of resistant bacteria.Furthermore,we also found that the overall up-regulation of nucleic acid synthesis is a new resistance mechanism of bacteria to combat to ciprofloxacin.Besides,the quantitative proteomics of whole cell proteins and lysine-crotonylated proteins combined with functional studies were used to investigate the mechanism of bacterial resistance to polymyxin.The results showed that bacteria might increase drug resistance by strengthening the modification,synthesis and transport of lipopolysaccharides,the synthesis of membrane phospholipids,the anti-acid mechanism,the phosphate metabolism,the efflux pump and porin,and etc.Meanwhile,lysine crotonylation might increase drug resistance of bacteria by participating in many biological processes,such as cell motility,energy synthesis,protein translation,nucleic acid synthesis,DNA replication,and amino acid metabolism.More importantly,through screening and validation,we found that berberine had more pronounced bactericidal effects on bacteria resistant to polymyxin than to other antibiotics.In conclusion,this study revealed the complex protein regulatory network in the mechanism of bacterial resistance by using quantitative proteomics technology and functional protein characterizations.From the perspective of time sequence,we found the evolutional directions of bacterial resistance,its continuity and stage features.The discovery of relevant mechanisms will provide a novel view for the development of antimicrobial drugs.