*4.2. AFM—Single Molecule Force Spectroscopy*

SMFS experiments were carried out on a Nanowizard II (JPK Instruments, Berlin, Germany) at room temperature. Prior to each experiment, every cantilever (Si3N4, Bruker MLCT-BIO, Cantilever D, Nominal spring constant k = 0.03 N/m) was individually calibrated using the Equipartition Theorem in the JPK software. Approximately 20 μL of protein (at a concentration of ~2 μM) were deposited onto an evaporated gold coverslip and allowed to adsorb for about 15 min. After this time, 1.8 mL of PBS buffer (pH 7.4, 150 mM) were added to reach an overall protein final concentration of ~20 nM. Constant-velocity, single-molecule pulling experiments were performed at 1 μm/s, with a recorded rate of 4096 Hz. Each experiment was carried out in fresh PBS buffer, to which EGCG (stock diluted in PBS) and DA (stock diluted in acidic MilliQ, pH 4) (stored at 4 ◦C protected from light) were added to reach the desired final concentration. Each solution was filtered on a filter screen with a porosity of 0.2 μm before each experiment.
