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| | NAD glycohydrolase assay |
| Description: | SmNACE activity and inhibition were measured with a sensitive, continuous, fluorometric assay platform that used 1,N6-ethenonicotinamide adenine dinucleotide (εNAD; Sigma-Aldrich, Inc.) as a substrate. This assay consists of measuring the appearance of the fluorescent εADP-ribose reaction product with excitation (λexc) and emission (λem) wavelengths of 310 and 410 nm, respectively, at 37 °C using the thermostatically controlled Spectramax Gemini XPS fluorescence plate reader (100 uL reaction volume; Molecular Devices, LLC) or the thermostatically controlled RF-5301PC spectrofluorophotometer (1 mL reaction volume; Shimadzu Scientific Instruments). The biochemical assay with 100 ug/mL of recombinant SmNACE was carried out in a 10 mM potassium phosphate buffer, at pH 7.4, containing 0.05% (w/v) emulphogen. |
| Affinity data for this assay | |
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