Fig. 3: PhLP2A modulation on the PFD-TRiC network.

a Atomic models of (i) PFD-bound TRiC (PDB:7WU7) and (ii) PhLP2A-bound TRiC. CCT3, CCT4, PFD and the PhLP2A NTD are highlighted. Contacts between TRiC and PFD or PhLP2A are indicated as colored-circles in the side view. Zoomed-in view of each contact between TRiC (CCT3, CCT4) and the cochaperones (PFD, PhLP2A). Negatively charged residues are indicated by red balls while positively charged residues are indicated by blue balls. Other residues are shown as stick cartoons. PFD and PhLP2A models are superimposed on their binding sites of CCT3 or CCT4. Red circles indicate the clash between the molecules. b (i) Schematic of the designed experiment. (ii, iii) 3D classification reveals PFD-bound TRiC (25.0%) and PhLP2A-bound TRiC (22.9%), respectively. The PFD-TRiC atomic model (PDB: 7WU7) fitted to the PFD-bound TRiC density. The red circle indicates no detectable PhLP2A density while the yellow circle indicates no detectable PFD density. c (i) Representative immunoblot of serially diluted PhLP2A (0.01–10 μM) bound to TRiC (0.25 μM). (ii) Quantification of immunoblots of dose dependent TRiC-bound PhLP2A establishes apparent dissociation constant. d (i) Native gel analysis using GFP-TRiC and cochaperones. Serially diluted PhLP2A (0.01–10 μM) were applied to the mixture of GFP-TRiC (0.25 μM) and AF-647 labeled PFD (1 μM). (ii) Serially diluted PFD (0.01–10 μM) were applied to the mixture of GFP-TRiC (0.25 μM) and AF-568 labeled PhLP2A (1 μM). (iii) Quantification of PhLP2A (orange) displaced from TRiC by PFD binding, and PFD (red) released as a result of PhLP2A binding. Source data are provided as a Source Data file.