Fig. 1: Conformations of ATL1_cyto dimers in the presence of GTPγS and GDP/AlF₄⁻ revealed by intramolecular smFRET assays.

a Ribbon representations of the crystal structures of ATL1_cyto dimers in Form 1 (left, PDB code 3QOF), Form 2 (middle, PDB code 3QNU), and Form 3 (right, PDB code 4IDN) conformations rendered in PyMOL. One protomer is shown in pink and the other in blue. The GTPase domains are shown in regular colors and the 3HBs in pale colors. The nucleotides are shown as orange sticks and the magnesium ions as yellow spheres. The selected T51/K400 pairs in one protomer for dye labeling are represented as green and red spheres, respectively. b Strategy of the intramolecular smFRET assays for ATL1_cyto-K-ATL1_cyto-TK dimers in the presence of GTPγS and GDP/AlF₄⁻. The dimer is formed by a C-terminal biotinylated ATL1_cyto-K and LD555/LD655-labeled ATL1_cyto-TK. The biotin-streptavidin interaction was used to immobilize the protein in a streptavidin-coated microfluidic chamber. c Representative fluorescence and smFRET trajectories of intramolecular smFRET assays for ATL1_cyto-K-ATL1_cyto-TK dimers in the presence of GTPγS (left) and GDP/AlF₄⁻ (right). LD555 (the donor) is shown in green, LD655 (the acceptor) in red, and FRET in dark blue. d Intramolecular smFRET distributions of ATL1_cyto-K-ATL1_cyto-TK dimers in the presence of GTPγS (left) and GDP/AlF₄⁻ (right). All of the individual FRET values with the number of molecules (Nm) displayed were compiled into a conformation-population histogram (gray lines) and fitted into a one-state GaussAmp distribution ( ~ 0.28). Each bar height represents the normalized count (%). The length of the error bar represents the normalized SD of a Poissom distribution from the count. Source data are provided as a Source Data file.