Table 1. Estimation of diffusion constants and bound/immobile fraction of EGFP-tagged wild-type (WT) and mutant (Y723F, N722S, W205G) Top1 from FRAP recovery studies.
| TOP1 variants | Treatment | Bound(immobile) fraction | Diffusion constant (μm2/s) |
|---|---|---|---|
| WT | No drug | 0.18 | 1.66 |
| CPT | 0.35 | 1.16 | |
| ABT | 0.22 | 0.79 | |
| ABT + CPT | 0.64 | 1.75 | |
| Y723F | No drug | 0.1 | 1.02 |
| CPT | 0.11 | 1.35 | |
| ABT | 0.1 | 1.31 | |
| ABT + CPT | 0.14 | 1.09 | |
| N722S | No drug | 0.18 | 1.21 |
| CPT | 0.17 | 1.37 | |
| ABT | 0.19 | 1.29 | |
| ABT + CPT | 0.19 | 1.46 | |
| W205G | No drug | 0.14 | 1.23 |
| CPT | 0.2 | 1.16 | |
| ABT | 0.22 | 1.12 | |
| ABT + CPT | 0.27 | 1.95 |
Normalized fluorescence intensities after photobleaching were fitted into the equation Equation 1 (see ‘Materials and Methods’ section for details) to quantitate bound/immobile fraction and diffusion constant of Top1 variants in the absence or presence Top1 poison (CPT), PARP inhibitor (ABT-888) and combination of ABT-888 + CPT. Quantification are based on mean FRAP data (n = 15).