R1.2 1 3 200 mesh copper grids

A 3 µL sample of protein solution [2.0 mg/mL PaFS, 50 mM HEPES (pH 7.5), 150 mM NaCl, 1.5 mM TCEP] was applied to glow-discharged (1 min, easiGlow, Pelco) R1.2/1.3 200-mesh copper grids (Quantifoil). Grids were blotted for 4.5 s at 100% humidity and flash-frozen with liquid ethane using a Vitrobot Mark I (FEI). Frozen grids were clipped and transferred to a Talos Arctica electron microscope (Thermo Fischer Scientific) operating at 200 keV (Pacific Northwest Cryo-EM Center). Images were recorded with a K2 direct electron detector at ×36,000 magnification (1.142 Å/pixel) at a nominal defocus of −1.5 to −2.7 µM, using SerialEM data collection software^{41 (link)}. 100-frame exposures were taken at 0.1 s/frame, using a dose rate of 5.6 e⁻/pixel/s (0.43 e⁻ Å⁻² per frame). A total of 1550 movies were recorded from one grid.

For the cryo-EM specimen preparation, R1.2/1.3 200 mesh copper grids (Quantifoil) were washed with ethyl acetate and glow-discharged by soft plasma ion bombardment (PIB-10, Vacuum Device Inc.). Aliquots (2.5 µL) of the p53-nucleosome complex or the p53 DBD-nucleosome complex were applied to the Quantifoil grids in the Vitrobot Mark IV chamber (Thermo Fisher Scientific) at 100% humidity and 16°C, and then blotted and plunged into liquid ethane. Data collections were performed on a Krios G4 cryo-transmission electron microscope (Thermo Fisher Scientific) operated at 300 kV, using the EPU software. Digital micrographs of the p53-nucleosome complex and the p53 DBD-nucleosome complex were recorded on a K3 BioQuantum (Gatan) direct electron detector calibrated at a pixel size of 1.06 and 1.1 Å in the electron counting mode, using a slit width of 25 eV, and retaining 40 frames with total doses of 56.8 and 46 electron/Ų, respectively (Table S1).