| Assay Method Information | |
| | Evaluation of Compounds as Inhibitors of GABA-AT |
| Description: | Inhibition constants were determined by monitoring GABA-AT activity in the presence of 0-50 mM concentrations of synthesized analogues using a coupled assay with the enzyme succinic semialdehyde dehydrogenase (SSDH). The assay solution consisted of 10 mM GABA, 5 mM α ketoglutarate, 1 mM NADP+, 5 mM β-mercaptoethanol, and excess SSDH in 50 mM potassium pyrophosphate buffer, pH 8.5. Enzyme activity was determined by observing the change in absorbance at 340 nm at 25° C. IC50 values were obtained using non-linear regression in GraphPad Prism5 software. Subsequent K1 values were determined using the Cheng-Prusoff relationship. (Yung-Chi, C.; Prusoff, W. H. Biochem. Pharmacol. 1973, 22, 3099-3108.) |
| Affinity data for this assay | |
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