Fig. 4: FICD’s TPR domain is essential for the recognition of ATP-bound unmodified BiP.

a The deAMPylation complex (coloured as in Fig. 1a with selected BiP interaction partners labelled) is aligned via its NBD (i) or SBDβ (ii) with an ADP-state BiP (PDB 7A4U; grey)⁷². Inset (i), a closeup view of FICD(TPR1)-BiP(NBD) contacts. (ii) the intermolecular β-sheet region of ℓ_7,8 (green) is shortened in BiP:ADP. Inset, disposition of Thr518 is highlighted (pink lines; hydrogen bonds). See Supplementary Movie 2 and Supplementary Fig. 6a. b FICD’s isolated TPR domain specifically binds the ATP-state of BiP. A representative BLI experiment demonstrating the ability of FICD(TPR) to engage immobilised BiP:ATP but not BiP:Apo. A global fit analysis of a one-phase association–dissociation model (black dashed lines) is overlaid. Below are the resulting kinetic binding parameters (mean ± SD) of the interaction of FICD(TPR) and BiP:ATP, from n = 3 independent experiments. c Steady-state equilibrium binding response analysis of the representative TPR domain binding experiment shown in b. Results from the analyte dilution series from the three independent BiP:ATP binding experiments are represented by different symbols. The fit from a one site binding model is shown with 95% confidence bands (dashed black line and solid grey lines, respectively). The calculated K_D is also annotated (mean ± SD). The inset panel highlights the same data and fitting over the lower analyte concentration range. Note, the lack of detectable steady-state binding of the TPR domain to BiP:Apo. d Representative BLI analysis of TPR domain mutants of monomeric (i) and dimeric (ii) FICD binding to immobilised ATP-bound BiP, from n = 3 independent experiments. See Supplementary Fig. 6b. Source data are provided as a Source Data file.