ID: ALA3888180
Type: Binding
Description: Homogenous Time-Resolved Fluorescence Assay (HTRF2 Assay): The standard assay conditions for the in vitro HTRF assay consisted of a 50 ul total reaction volume in black 384-well Costar polypropylene plates in 1xPBS buffer pH 7.4, 1 mM DTT, 0.1% BSA, 0.2 nM GST-hMDM2 (aa 1-188), 0.5 nM biotinylated-p53 (aa 1-83), 0.18 nM SA-XLent (Cisbio; Bedford, Mass.), 0.6 nM anti-GST cryptate monoclonal antibody (Cisbio; Bedford, Mass.) and 100 mM KF. Amino acid residues 1-188 of human MDM2 were expressed as an amino-terminal glutathione-5-transferase (GST) fusion protein (GST-hMDM2) in Escherichia coli. Residues 1-83 of human p53 were expressed as an amino-terminal AviTag-TrxA-6×His fusion protein (biotinylated p53) in E. coli. Each protein was purified from cell paste by affinity chromatography. Specifically, 10 uL of GST-hMDM2 was incubated with 10 ul of diluted compound (various concentrations, serially diluted) in 10% DMSO for 20 minutes at room temperature. 20 uL of biotinylated-p53 was added to the GST-hMDM2+compound mixture, and then incubated at room temperature for 60 min. 10 uL of detection buffer consisting of SA-XLent, anti-GST cryptate antibody and KF was added to GST-hMDM2, biotinylated-p53 and compound reaction and left at room temperature to reach equilibrium for >4 hrs. The final concentration of DMSO in the reaction was 2%. Time-resolved fluorescence readings were measured on a microplate multilabel reader. Percentage of inhibition was calculated relative to nutlin-3.
Bioactivity
Activity Types for Assay ALA3888180
IC50
Parent Molecular Weight | ALogP | Polar Surface Area |
---|---|---|
478.42 | 5.69 | 5.69 |
558.51 | 5.93 | 5.93 |
464.39 | 5.30 | 5.30 |
583.58 | 6.03 | 6.03 |
597.61 | 6.37 | 6.37 |
593.57 | 5.73 | 5.73 |
595.59 | 5.98 | 5.98 |
567.54 | 5.20 | 5.20 |
579.55 | 5.35 | 5.35 |
593.57 | 5.74 | 5.74 |
526.48 | 5.35 | 5.35 |
540.51 | 5.74 | 5.74 |
552.52 | 5.89 | 5.89 |
554.54 | 6.13 | 6.13 |
568.56 | 6.52 | 6.52 |
566.55 | 6.14 | 6.14 |
566.55 | 6.28 | 6.28 |
580.57 | 6.67 | 6.67 |
568.52 | 5.12 | 5.12 |
582.59 | 6.77 | 6.77 |
580.57 | 6.52 | 6.52 |
582.59 | 6.77 | 6.77 |
566.55 | 6.13 | 6.13 |
540.51 | 5.60 | 5.60 |
538.49 | 5.35 | 5.35 |
568.56 | 6.38 | 6.38 |
492.44 | 6.08 | 6.08 |
520.50 | 6.86 | 6.86 |
541.50 | 4.86 | 4.86 |
553.51 | 4.86 | 4.86 |
581.56 | 5.64 | 5.64 |
607.60 | 6.13 | 6.13 |
609.57 | 4.97 | 4.97 |
511.45 | 7.08 | 7.08 |
511.45 | 7.08 | 7.08 |
525.48 | 7.32 | 7.32 |
545.89 | 7.73 | 7.73 |
434.36 | 5.94 | 5.94 |
460.40 | 6.33 | 6.33 |
617.60 | 6.60 | 6.60 |
567.54 | 5.39 | 5.39 |
649.04 | 7.83 | 7.83 |
546.41 | 6.62 | 6.62 |
581.56 | 5.73 | 5.73 |
596.57 | 5.90 | 5.90 |
533.50 | 5.64 | 5.64 |
564.53 | 5.89 | 5.89 |
512.44 | 6.47 | 6.47 |
664.70 | 6.20 | 6.20 |
600.59 | 5.86 | 5.86 |
581.56 | 5.78 | 5.78 |
600.54 | 5.98 | 5.98 |
651.70 | 5.33 | 5.33 |
558.50 | 5.88 | 5.88 |
653.00 | 7.58 | 7.58 |
606.49 | 6.29 | 6.29 |
628.58 | 6.16 | 6.16 |
578.56 | 6.14 | 6.14 |
531.48 | 5.93 | 5.93 |
518.44 | 6.02 | 6.02 |
638.01 | 6.94 | 6.94 |
534.52 | 7.25 | 7.25 |
586.55 | 6.66 | 6.66 |
610.60 | 4.89 | 4.89 |
587.47 | 6.72 | 6.72 |
643.63 | 7.25 | 7.25 |
580.57 | 6.67 | 6.67 |
654.66 | 4.12 | 4.12 |
572.53 | 6.27 | 6.27 |
628.58 | 6.16 | 6.16 |
580.57 | 6.53 | 6.53 |
596.57 | 5.76 | 5.76 |
552.52 | 5.74 | 5.74 |
568.52 | 4.79 | 4.79 |
580.57 | 6.53 | 6.53 |
633.59 | 6.30 | 6.30 |
638.01 | 6.94 | 6.94 |
632.58 | 7.31 | 7.31 |
554.54 | 6.14 | 6.14 |
567.54 | 5.34 | 5.34 |
616.61 | 6.21 | 6.21 |
538.54 | 6.47 | 6.47 |
518.48 | 6.88 | 6.88 |
594.60 | 6.91 | 6.91 |
506.43 | 5.87 | 5.87 |
545.51 | 5.78 | 5.78 |
604.56 | 5.68 | 5.68 |
541.48 | 6.17 | 6.17 |
460.40 | 6.47 | 6.47 |
636.54 | 7.06 | 7.06 |
620.64 | 7.45 | 7.45 |
518.48 | 6.74 | 6.74 |
541.50 | 5.14 | 5.14 |
630.64 | 6.52 | 6.52 |
553.51 | 5.28 | 5.28 |
560.44 | 7.01 | 7.01 |
579.45 | 8.10 | 8.10 |
583.53 | 4.63 | 4.63 |
544.47 | 5.49 | 5.49 |
595.59 | 6.12 | 6.12 |
614.59 | 7.09 | 7.09 |
595.59 | 6.12 | 6.12 |
569.55 | 5.92 | 5.92 |
494.46 | 4.96 | 4.96 |
573.51 | 5.39 | 5.39 |
624.63 | 4.76 | 4.76 |
561.55 | 6.42 | 6.42 |
608.63 | 7.16 | 7.16 |
568.56 | 6.38 | 6.38 |
594.60 | 6.92 | 6.92 |
568.56 | 6.52 | 6.52 |
581.56 | 5.59 | 5.59 |
595.59 | 6.12 | 6.12 |
582.55 | 5.13 | 5.13 |
595.47 | 5.75 | 5.75 |
616.61 | 6.21 | 6.21 |
597.61 | 6.37 | 6.37 |
597.56 | 5.11 | 5.11 |
562.41 | 5.94 | 5.94 |
597.56 | 4.88 | 4.88 |
610.60 | 5.33 | 5.33 |
618.55 | 6.92 | 6.92 |
539.45 | 6.26 | 6.26 |
609.47 | 7.19 | 7.19 |
632.58 | 7.31 | 7.31 |
582.59 | 6.91 | 6.91 |
492.44 | 6.08 | 6.08 |
578.56 | 6.28 | 6.28 |
555.52 | 5.20 | 5.20 |
490.43 | 6.29 | 6.29 |
569.55 | 5.64 | 5.64 |
580.57 | 6.52 | 6.52 |
608.51 | 6.53 | 6.53 |
568.56 | 6.52 | 6.52 |
554.54 | 6.13 | 6.13 |
584.56 | 5.50 | 5.50 |
434.36 | 5.72 | 5.72 |
490.43 | 5.55 | 5.55 |
614.59 | 7.17 | 7.17 |
592.59 | 6.67 | 6.67 |
630.64 | 6.52 | 6.52 |
609.62 | 6.51 | 6.51 |
478.42 | 5.69 | 5.69 |
617.60 | 6.60 | 6.60 |
566.55 | 6.28 | 6.28 |
581.56 | 5.78 | 5.78 |
623.52 | 6.13 | 6.13 |
592.52 | 6.65 | 6.65 |
581.56 | 5.73 | 5.73 |
547.52 | 6.03 | 6.03 |
581.56 | 5.04 | 5.04 |
629.55 | 6.33 | 6.33 |
573.52 | 5.09 | 5.09 |
534.48 | 6.65 | 6.65 |
587.47 | 5.92 | 5.92 |
617.54 | 6.18 | 6.18 |
587.54 | 5.73 | 5.73 |
533.45 | 5.76 | 5.76 |
574.55 | 5.47 | 5.47 |
668.68 | 4.51 | 4.51 |
494.46 | 4.96 | 4.96 |
638.66 | 5.15 | 5.15 |
605.59 | 5.25 | 5.25 |
635.01 | 7.44 | 7.44 |
606.61 | 7.06 | 7.06 |
554.54 | 6.14 | 6.14 |
578.56 | 6.28 | 6.28 |
520.50 | 6.72 | 6.72 |
600.56 | 6.78 | 6.78 |
602.58 | 5.82 | 5.82 |
586.52 | 5.64 | 5.64 |
570.51 | 6.03 | 6.03 |
636.64 | 5.42 | 5.42 |
654.66 | 4.12 | 4.12 |
603.57 | 6.29 | 6.29 |
601.55 | 6.18 | 6.18 |
490.43 | 5.69 | 5.69 |
615.58 | 6.51 | 6.51 |
616.61 | 6.13 | 6.13 |
574.47 | 7.40 | 7.40 |
594.60 | 6.91 | 6.91 |
533.50 | 5.64 | 5.64 |
574.47 | 7.40 | 7.40 |
611.59 | 5.27 | 5.27 |
596.58 | 5.16 | 5.16 |
561.55 | 6.42 | 6.42 |
478.42 | 5.69 | 5.69 |
604.56 | 5.68 | 5.68 |
595.59 | 5.98 | 5.98 |
635.01 | 7.44 | 7.44 |
492.44 | 5.94 | 5.94 |
568.56 | 6.52 | 6.52 |
680.70 | 5.44 | 5.44 |
580.57 | 6.53 | 6.53 |
608.58 | 5.76 | 5.76 |
632.58 | 7.31 | 7.31 |
552.52 | 5.74 | 5.74 |
567.54 | 5.25 | 5.25 |
654.62 | 7.33 | 7.33 |
607.60 | 6.12 | 6.12 |
602.58 | 5.82 | 5.82 |
518.48 | 6.93 | 6.93 |
630.59 | 5.82 | 5.82 |
581.56 | 5.64 | 5.64 |
595.54 | 5.26 | 5.26 |
533.50 | 6.22 | 6.22 |
547.52 | 6.47 | 6.47 |
654.62 | 7.33 | 7.33 |
518.44 | 5.87 | 5.87 |
476.40 | 5.30 | 5.30 |
581.56 | 5.64 | 5.64 |
605.58 | 5.78 | 5.78 |
592.46 | 6.24 | 6.24 |
554.54 | 5.99 | 5.99 |
658.67 | 4.73 | 4.73 |
515.48 | 7.25 | 7.25 |
492.44 | 5.94 | 5.94 |
523.46 | 7.08 | 7.08 |
569.55 | 5.50 | 5.50 |
621.63 | 6.56 | 6.56 |
727.78 | 4.24 | 4.24 |
596.58 | 5.05 | 5.05 |
629.61 | 6.78 | 6.78 |
463.41 | 5.79 | 5.79 |
486.45 | 6.00 | 6.00 |
670.08 | 7.35 | 7.35 |
554.54 | 5.99 | 5.99 |
553.51 | 4.86 | 4.86 |
630.59 | 6.17 | 6.17 |
486.44 | 6.72 | 6.72 |
504.45 | 6.08 | 6.08 |
434.36 | 5.72 | 5.72 |
616.61 | 6.13 | 6.13 |
473.40 | 6.22 | 6.22 |
554.54 | 6.14 | 6.14 |
494.42 | 4.66 | 4.66 |
476.40 | 5.90 | 5.90 |
583.58 | 6.03 | 6.03 |
566.55 | 5.22 | 5.22 |
569.53 | 7.31 | 7.31 |
630.59 | 6.17 | 6.17 |
566.55 | 6.14 | 6.14 |
661.62 | 7.31 | 7.31 |
452.35 | 6.08 | 6.08 |
490.43 | 5.69 | 5.69 |
520.45 | 6.26 | 6.26 |
462.42 | 6.72 | 6.72 |
656.05 | 7.00 | 7.00 |
643.63 | 7.17 | 7.17 |
593.57 | 5.78 | 5.78 |
621.61 | 6.35 | 6.35 |
673.66 | 6.78 | 6.78 |
535.47 | 6.05 | 6.05 |
607.60 | 6.17 | 6.17 |
649.66 | 6.83 | 6.83 |
655.64 | 7.31 | 7.31 |
434.38 | 7.18 | 7.18 |
581.56 | 5.64 | 5.64 |
661.62 | 7.31 | 7.31 |
478.42 | 5.69 | 5.69 |
595.59 | 6.03 | 6.03 |
540.51 | 5.74 | 5.74 |
647.60 | 6.92 | 6.92 |
594.60 | 6.91 | 6.91 |
554.54 | 6.13 | 6.13 |
581.56 | 5.78 | 5.78 |
463.40 | 6.12 | 6.12 |
645.60 | 6.96 | 6.96 |
432.35 | 5.55 | 5.55 |
492.40 | 5.48 | 5.48 |
581.56 | 5.64 | 5.64 |
595.59 | 6.03 | 6.03 |
532.46 | 6.26 | 6.26 |
582.55 | 4.71 | 4.71 |
579.55 | 5.34 | 5.34 |
593.57 | 5.73 | 5.73 |
630.59 | 5.82 | 5.82 |
504.45 | 6.49 | 6.49 |
568.56 | 6.38 | 6.38 |
635.64 | 6.69 | 6.69 |
567.54 | 5.35 | 5.35 |
629.61 | 6.43 | 6.43 |