New Delhi Metallo-β-lactamase (NDM) grants resistance to a broad spectrum of β-lactam antibiotics including last-resort carbapenems and is emerging as a global antibiotic resistance threat. Limited zinc availability adversely impacts the ability of NDM-1 to provide resistance, but a number of clinical variants have emerged that are more resistant to zinc scarcity (e.g., NDM-15). To provide a novel tool to better study metal ion sequestration in host-pathogen interactions, we describe the development of a fluorescent probe that reports on the dynamic metallation state of NDM within E. coli. The thiol-containing probe selectively coordinates the dizinc metal cluster of NDM and results in a 17-fold increase in fluorescence intensity. Reversible binding enables competition and time-dependent studies that reveal fluorescence changes used to detect enzyme localization, substrate and inhibitor engagement, and changes to metallation state through the imaging of live E. coli using confocal microscopy. NDM-1 is shown to be susceptible to demetallation by intracellular and extracellular metal chelators in a live-cell model of zinc dyshomeostasis, whereas the NDM-15 metallation state is shown to be more resistant to zinc flux. The development of this reversible turn-on fluorescent probe for the metallation state of NDM provides a new tool for monitoring the impact of metal ion sequestration by host defense mechanisms and to detect inhibitor target engagement during the development of therapeutics to counter this resistance determinant.

We report the synthesis and application of a small molecule probe for carbonic anhydrase (CA) to track holo-CA in cell lysates and live-cell models of zinc dyshomeostasis. The probe displays a 12-fold increase in fluorescence upon binding to bovine CA and also responds to human CA isoforms.

A monotyrosine based copolymer was synthesized, characterized and studied for its interaction with DNA for potential biological applications.