The effects of NDM-5 on and the screening of interacting proteins.

Carbapenem-resistant () strains are widely distributed and spreading rapidly, creating significant challenges for clinical therapeutics. NDM-5, a novel mutant of New Delhi Metallo-β-Lactamase-1 (NDM-1), exhibits high hydrolase activity toward carbapenems. Since the genetic backgrounds of clinically isolated carbapenem-resistant are heterogeneous, it is difficult to accurately evaluate the impact of on antibiotic resistance. Herein, BL21 was transformed with a plasmid harboring , and the resultant strain was named BL21 (pET-28a-). Consistent with the findings of previous studies, the introduction of exogenous resulted in markedly greater resistance of to multiple β-lactam antibiotics. Compared with BL21 (pET-28a), BL21 (pET-28a-) exhibited reduced motility but a significant increase in biofilm formation capacity. Furthermore, transcriptome sequencing was conducted to compare the transcriptional differences between BL21 (pET-28a) and BL21 (pET-28a-). A total of 461 differentially expressed genes were identified, including those related to antibiotic resistance, such as genes associated with the active efflux system (, and ), pili (, and ), biofilm formation (, and ) and antioxidant processes (). Finally, the pGS21a plasmid harboring was transformed into Rosetta2, after which the expression of the NDM-5 protein was induced using isopropyl-β-D-thiogalactoside (IPTG). Using glutathione-S-transferase (GST) pull-down assays, total proteins from were scanned to screen out 82 proteins that potentially interacted with NDM-5. Our findings provide new insight into the identified proteins to identify potential antibiotic targets and design novel inhibitors of carbapenem-resistant bacteria.