Fig. 4: XL-MS reveals the binding interface of trigger factor–GAPDH complex.

a SDS–PAGE analysis of the disuccinimidyl dibutyric urea cross-linked species shows a homogeneous cross-linked trigger factor–GAPDH complex. Note that above the band corresponding to the crosslinked complex there are additional crosslinked species corresponding to higher order oligomeric assemblies, but these represent only a small proportion of the crosslinked products. These experiments were independently repeated three times with similar results. b The intermolecular cross-links found in the trigger factor–GAPDH complex are shown as teal solid lines. All the lysine residues across both protein sequences are marked in teal. Three structural domains of trigger factor, including the N-terminal domain (NTD), Peptidyl-prolyl isomerase domain (PPIase domain) and the C-terminal domain (CTD), are labelled. A list of all intermolecular crosslinked peptides can be found in Supplementary Table 1. c The cross-linked sites, shown as green spheres, are labelled and mapped onto the crystal structure of trigger factor (pdb: 1w26). Non-cross-linked lysine residues are shown as sticks. The three structural domains of trigger factor are labelled as outlined in b. d The cross-linked sites, shown as pale-yellow spheres, are labelled and mapped onto a subunit of crystal structure of rabbit-muscle GAPDH (pdb: 1J0X). The oligomeric interface of native GAPDH is coloured in pink. The two structural domains of GAPDH, catalytic domain and cofactor-binding domain, are labelled. Non-cross-linked lysine residues are shown as sticks.