The PDF file includes:

• Fig. S1. Structure and conservation of TcAs.
• Fig. S2. Cryo-EM of Pl-TcdA1, Pl-TcdA4, and Xn-XptA1.
• Fig. S3. Cryo-EM of Mm-TcdA4 and Yp-TcaATcaB and purification of Yp-TcaATcaB(WT) and Yp-TcaATcaB–∆622–714.
• Fig. S4. A 3₁ trefoil protein knot is present in all five TcAs.
• Fig. S5. Biophysical properties of the TcA channels.
• Fig. S6. pH stability of Yp-TcaATcaB and characterization of chimeric holotoxin formation.
• Fig. S7. Intoxication of HEK293T cells with chimeric holotoxins.
• Fig. S8. Topology of neuraminidase-like domain and nonconserved cluster of three histidine residues.
• Fig. S9. pH-induced pore formation of Xn-XptA1, Mm-TcdA4, and Yp-TcaATcaB.
• Fig. S10. Mutational studies of Pl-TcdA1.
• Legends for movies S1 and S2

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Other Supplementary Material for this manuscript includes the following:

• Movie S1 (.mov format). Cryo-EM density maps of Pl-TcdA1, Pl-TcdA4, Xn-XptA1, Mm-TcdA4, and Yp-TcaATcaB.
• Movie S2 (.mov format). Molecular trefoil knot in Pl-TcdA1.