Fig. 1: Structural characterization of the UFC1-UBS interaction.

a Schematic representation of UBA5 elements. UIS (UFM1-interacting motif); UBS (UFC1-binding sequence) b Trans-binding mechanism of UFM1 and UFC1 to the homodimeric UBA5. UFM1 and UFC1 bind the UIS and UBS, respectively, of one molecule of UBA5 and interacts with the adenylation domain of the other UBA5 molecule. For simplicity, the UIS and UBS are shown only for one molecule of UBA5. The black arrow indicates the linker connecting the UIS to UBS; AD-adenylation domain. c Crystal structure of UFC1 (brown) bound to UBS (red); black arrow points on the active site Cys116 of UFC1 (yellow sphere). d Contact between UBS and UFC1 α-helix 1 and β-strand 1. Sidechains of UFC1 residues involved in UBS binding are shown in stick representation. e M401 (highlighted by yellow arrow) of UBS (red) occupies a hydrophobic pocket on the surface of UFC1 (brown) f SDS-PAGE analysis showing the effect of UBS mutations on UFM1 transfer to UFC1. The gel is representative of two independent experiments.