| Solution Information | help | |
| Enzyme: | Mu-type opioid receptor | |
| inhibitor: | BDBM64868 | |
| 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 the PP1 inhibition potency of compounds identified as active in a previous set of experiments entitled, "Counterscreen for inhibitors of PP5: fluorescence-based biochemical high throughput primary assay to identify inhibitors of Protein Phosphatase 1 (PP1)" (AID 2235). This biochemical, fluorescence-based assay employs purified PP1alpha (catalytic subunit) and 3-O-methylfluorescein phosphate (OMFP) as the substrate. Cleavage of OMFP by PP1alpha converts OMFP to the fluorescent product 3-O-methylfluorescein. As designed, compounds that act as PP1alpha inhibitors will block PP1alpha catalytic activity, thereby decreasing OMFP cleavage and production of fluorescent product, resulting in reduced well fluorescence. Compounds are tested in triplicate in a 10-point 1:3 dilution series starting at a nominal test concentration of 67 micromolar. Protocol Summary: Prior to the start of the assay, 3 microliters of assay buffer (40 mM HE | |
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