AUTHOR=Li Lin , Gao Yiming , Wang Longbo , Lu Fang , Ji Qianyu , Zhang Yanfang , Yang Shuo , Cheng Ping , Sun Feifei , Qu Shaoqi 

TITLE=The effects of NDM-5 on Escherichia coli and the screening of interacting proteins

JOURNAL=Frontiers in Microbiology

VOLUME=15

YEAR=2024

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2024.1328572

DOI=10.3389/fmicb.2024.1328572

ISSN=1664-302X

ABSTRACT=<p>Carbapenem-resistant <italic>Escherichia coli</italic> (<italic>E. coli</italic>) 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 <italic>E. coli</italic> are heterogeneous, it is difficult to accurately evaluate the impact of <italic>bla</italic><sub>NDM–5</sub> on antibiotic resistance. Herein, <italic>E. coli</italic> BL21 was transformed with a plasmid harboring <italic>bla</italic><sub>NDM–5</sub>, and the resultant strain was named BL21 (pET-28a-<italic>bla</italic><sub>NDM–5</sub>). Consistent with the findings of previous studies, the introduction of exogenous <italic>bla</italic><sub>NDM–5</sub> resulted in markedly greater resistance of <italic>E. coli</italic> to multiple β-lactam antibiotics. Compared with BL21 (pET-28a), BL21 (pET-28a-<italic>bla</italic><sub>NDM–5</sub>) 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-<italic>bla</italic><sub>NDM–5</sub>). A total of 461 differentially expressed genes were identified, including those related to antibiotic resistance, such as genes associated with the active efflux system (<italic>yddA</italic>, <italic>mcbR</italic> and <italic>emrY</italic>), pili (<italic>csgC</italic>, <italic>csgF</italic> and <italic>fimD</italic>), biofilm formation (<italic>csgD</italic>, <italic>csgB</italic> and <italic>ecpR</italic>) and antioxidant processes (<italic>nuoG</italic>). Finally, the pGS21a plasmid harboring <italic>bla</italic><sub>NDM–5</sub> was transformed into <italic>E. coli</italic> 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 <italic>E. coli</italic> 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.</p>