Ebsulfur as a potent scaffold for inhibition and labelling of New Delhi metallo-β-lactamase-1 in vitro and in vivo
The superbug infection caused by New Delhi metallo-β-lactamase (NDM-1) has grown into an emerging threat, labelling and inhibition of NDM-1 has proven challenging due to its shuttling between pathogenic bacteria. Here, we report a potent covalent scaffold, ebsulfur, for targeting the protein in vitro and in vivo . Enzymatic kinetic study indicated that eighteen ebsulfurs gained except 1a – b and 1f inhibited NDM-1, exhibiting an IC 50 value ranging of 0.16–9 μM, and 1g was found to be the best, dose- and time-dependent inhibitor with an IC 50 of 0.16 μM. Also, these ebsulfurs effectively restored the antibacterial activity of cefazolin against E. coli expressing NDM-1, and the best effect was observed to be from 1g , 1i and 1n , resulting in an 256-fold reduction in MIC of the antibiotic at a dose of 16 μg/mL. The equilibrium dialysis study implied that the ebsulfur disrupted the coordination of one Zn(II) ion at active site of NDM-1. Labelling of NDM-1 using a constructed fluorescent ebsulfur Ebs - R suggested that the inhibitor covalently bound to the target through SDS-PAGE analysis in vitro . Also, labelling NDM-1 in living E. coli cells with Ebs - R by confocal microscopic imaging showed the real-time distribution change process of intracellular recombinant protein NDM-1. Moreover, the cytotoxicity of these ebsulfurs against L929 mouse fibroblastic cells was tested, and their capability to restore antibacterial activity of antibiotic against clinical strains E. coli EC08 producing NDM-1 was determined. The ebsulfur scaffold proposed here is valuable for development of the covalent irreversible inhibitors of NDM-1, and also for labelling the target in vitro and in vivo .
危险品化学品经营许可证（带存储）