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Assay Method Information |
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| | Radioactive Assay |
| Description: | To measure the kinetic constants by radioactive assays, 0.2 μM wild-type E. coli OPPS and the wild-type or mutant S. cerevisiae GGPPS and human GGPPS were mixed with various concentrations of either FPP or MANT-O-GPP, and [14C]IPP as specified. For FPP or MANT-O-GPP Km measurements, 0.5−5 μM FPP or MANT-O-GPP was used with 50 μM [14C]IPP. For [14C]IPP Km and kcat determinations, 3.5 μM FPP or MANT-O-GPP was utilized to saturate the enzyme and 2−50 μM [14C]IPP was added; 40 μL portions of the reaction mixtures were periodically withdrawn and mixed with 10 mM EDTA to chelate with Mg2+ and stop the enzyme reactions. |
| Affinity data for this assay | |
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