Fig. 1: Selection of high affinity, STAT3-selective monobody binders.

a Schematic representation of the recombinant STAT3 constructs used for monobody selection. b Binding titration of yeast cells displaying monobodies (MS3-6: light blue line, MS3-N3: dark blue line) at their surface to recombinant STAT3 proteins. The mean fluorescence intensity of yeast cells bound to the target are plotted as a function of the protein concentration. Data from three individual experiments, mean ± SD are shown and a curve fitting 1:1 binding model was used. c Isothermal calorimetric titration (ITC) of STAT3 (100 µM) to a MS3-6 solution (10 µM) performed at 25 °C. The upper panel shows raw heat signal, while the lower panel shows the integrated calorimetric data of the area for each peak. A best fit 1:1 binding model was used and is illustrated by a black line (Microcal software). K_D and stoichiometry values (N) are indicated in the figure. ∆H (kcal mol⁻¹) = −22.5 ± 0.5; ∆G (kcal mol⁻¹) = −11.1. d Representative immunoblot analysis from three independent experiments of monobody-VHL fusion expression overtime upon doxycycline treatment (1 µg ml⁻¹) leading to STAT3 degradation. e STAT3 degradation levels were quantified and plotted from three independent experiments. Mean ± SD are shown and significance is indicated according to a two-sided unpaired t-test against HA4-Y87A control: MS3-6 *P = 0.033, **P = 0.0011, MS3-N3 *P = 0.0125. Source data are provided as a source data file.