In total, 3 μL of protein sample at 3 mg/mL (added with 0.025% DDM) was applied onto a H2/O2 glow-discharged, 200-mesh Quantifoil R0.6/1.0 grid (Quantifoil, Micro Tools GmbH, Germany). The grid was then blotted for 3.0 s with a blot force of 0 at 8°C and 100% humidity and plunge-frozen in liquid ethane using a Vitrobot (Thermo Fisher Scientific, USA). Cryo-EM data were collected with a 300 keV Titan Krios electron microscope (Thermo Fisher Scientific, USA) and a K3 direct electron detector (Gatan, USA). Images were recorded at 22500 × magnification and calibrated at a super-resolution pixel size of 0.82 Å/pixel. The exposure time was set to 2 s with a total accumulated dose of 60 electrons per Å2. All images were automatically recorded using SerialEM. A total of 12,704 images were collected with a defocus range from −2.0 μm to −1.0 μm. Statistics for data collection and refinement are in Table S1. The methods for processing are described in Figures S2 and andS4S4.

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