Standard pulse sequence hsqcetfpf3gpsi

All NMR titration experiments were performed at 298 K on a Bruker 900 MHz spectrometer equipped with a cryogenic probe in an NMR buffer of 50 mM HEPES (pH 7.0), 50 mM NaCl, and 10% D2O. Each NMR sample was made with a volume of 500 μL, containing 10 μM 15 N labeled TDP-43 LCD. We performed the titration experiment by addition of 5 μM or 10 μM HSPB1 and it's variant respectively. Bruker standard pulse sequence (hsqcetfpf3gpsi) was used to collect the 2D 1 H- 15 N HSQC spectrum with 32 scans, and 2048 × 160 complex points were used for 1 H (14 ppm) and 15 N (21 ppm) dimension, respectively. Backbone assignment of TDP-43 LCD was accomplished according to the previous publication 20 . All NMR data were processed by NMRPipe 128 and analyzed by Sparky 129 .

Backbone assignment of TDP-43 LCD was accomplished according to the previous publication (Conicella et al., 2016) . All NMR titration experiments were performed at 298 K on a Bruker 900 MHz spectrometer equipped with cryogeni probe in an NMR buffer of 20 mM MES (pH 6.0), 150 mM NaCl, 10% glycerol and 20% D 2 O. Here, we increased salt concentration and utilized glycerol to avoid LLPS of TDP-43 LCD with Hsp70. Each NMR sample was made with a volume of 500 μL, containing 15 N-TDP-43 LCD (20 μM) desalted from denature buffer freshly with/without Hsp70 WT and its variants as indicated. Bruker standard pulse sequence (hsqcetfpf3gpsi) was used to collect the 2D 1 H-15 N HSQC spectrum with 16 scans. And 2048 × 160 complex points were used for 1 H (14 ppm) and 15 N (21 ppm) dimension, respectively. All NMR data were processed by NMRpipe and analysed by SPARK (Delaglio et al., 1995; Lee et al., 2015) .