| Solution Information | help | |
| Enzyme: | Beta-lactamase | |
| inhibitor: | BDBM51998 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | Assay Overview: The purpose of this assay is to determine dose response curves for compounds identified as active in a previous set of experiments entitled, "Primary biochemical high throughput screening assay to identify inhibitors of VIM-2 metallo-beta-lactamase" (PubChem AID 1527), and inactive in a set of experiments entitled, "Epi-absorbance primary biochemical high throughput screening assay to identify inhibitors of IMP-1 metallo-beta-lactamase" (PubChem AID 1556). This assay also serves as an orthogonal counterscreen to identify epi-absorbance assay artifacts. This biochemical FRET-based assay employs as the beta-lactamase substrate the cephalosporin scaffold CCF2 that has been tagged with a donor (7-hydroxycoumarin) and quencher/acceptor (fluorescein) FRET pair. VIM-2 mediated hydrolysis of the CCF2 increases donor fluorescence at 460nm and decreases acceptor fluorescence at 535nm. In this assay, test compounds are incubated with purified VIM-2 enzyme and CCF2 in detergent-c | |
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