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Details of RareLED ID Assay_1028
Primary information
GENEGNPTAB
NUMBER OF ASSAY9
ASSAY IDs568345 299525 1258558 1236209 1236173 1236172 1324705 1324701 1324706
Assay Name1. Inhibition of human OGT,2. Binding affinity to human OGT expressed in Escherichia coli BL1 (DE3) cells by azido-ELISA,3. Inhibition Assay: 384-well plates (Costar #3654) were filled using a liquid handling robot with 20 uL of a mixture of 50 nM of a fluorescein-linked UDP-GlcNAc analog (see Gross et al, 2003), 1-2 uM sOGT, and buffer (20 mM potassium phosphate, pH=7.4 with 500 uM tris(hydroxypropyl)phosphine). The 1249 compound library was serially diluted in DMSO from the 5 mg/ml plates fivefold 3 times, such that 4 different concentrations of compounds were prepared. Compound libraries of the 4 concentrations in duplicated were then transferred to the assay plates using a 100 nL pin array, resulting in a final compound concentration of 25 ug/mL or 70 uM at the highest of the four concentrations, assuming an average compound MW of 350. Using a Perkin Elmer Envision microplate reader, the sample was excited at 480 nm in the vertical plane, and simultaneous emission intensity (535 nm) of the vertical and horizontal polarization planes was measured.,4. Inhibition of nucleocytoplasmic OGT (unknown origin),5. Inhibition of human OGT using [3H]-UDP-GlcNAc as donor and recombinant nup2 as acceptor by Dixon plot analysis,6. Inhibition of human OGT using [3H]-UDP-GlcNAc as donor and recombinant nup2 as acceptor by liquid scintillation counting analysis,7. Noncompetitive inhibition of OGT (unknown origin) using variable UDP-GlcNAc levels and fixed CKII peptide level by UDP-Glo glycosyltransferase assay,8. Inhibition of OGT (unknown origin) assessed as reduction in GlcNAc transfer to CKII peptide incubated for 60 mins by HPLC method,9. Noncompetitive inhibition of OGT (unknown origin) using fixed UDP-GlcNAc levels and variable CKII peptide level by UDP-Glo glycosyltransferase assay
ACTIVITYSubstance BioActivity: 1 Active, 1 Tested Substance BioActivity: 3 Active, 2 Activity <= 1 µM, 3 Tested Substance BioActivity: 20 Active, 1 Activity <= 1 µM, 26 Tested Substance BioActivity: 1 Active, 3 Tested Substance BioActivity: 1 Active, 3 Tested Substance BioActivity: 1 Active, 6 Tested Substance BioActivity: 1 Active, 2 Tested Substance BioActivity: 1 Active, 13 Tested Substance BioActivity: 2 Tested