Extended Data Fig. 6: The N-terminal region of ARF1 is required for SopF catalytic activation.
a, Multiple sequence alignment of the ARF family. Secondary structures of ARF1 are marked and labeled along the sequence. The alignment was performed using the Clustal Omega algorithm. Identical residues are highlighted by red background and conserved residues are in red. The N-terminal and switch I/II regions of ARFs are highlighted by pink and green boxes, respectively. Residues in ARF1 that make contacts with SopF in are marked by blue background. The residue number is indicated on the left of the sequence. b–d, NAD+-hydrolysis activity of SopF ∆N62 in complex with different ARFs. The experiments were performed similarly as in Fig. 3d. Kinetic parameters of NAD+ hydrolysis by the SopF–ARF complexes were determined. d, ARF1N16-ARF5Q71L ΔN16 was generated by replacing the N-terminal 16 residues in ARF5 (Q71L) with that of ARF1. e, ADP-ribosylation of ATP6V0C by indicated SopF–ARF complexes in the reconstitution system. Data in b, d are mean values ± SD from three determinations. f. Structural comparison of NAD+-bound SopF–ARF1 complex and cholera toxin subunit A1 (CTA1) in complex with ARF6 (PDB code: 2A5F). The structure of ARF6 in CTA1–ARF6 complex was superimposed with that of ARF1 in the SopF–ARF1 complex. The switch I/II of ARF1/6 are highlighted with different colors as indicated. The bound NAD+ is shown as ball and sticks models. The N-terminus of ARFs are marked. Data (b–e) are representative of two independent experiments.
