Extended Data Fig. 2: BB-loop and AE interface are important for phase separation of TIR ___domain proteins.
From: Substrate-induced condensation activates plant TIR ___domain proteins
a, A schematic of the enzymatic active TIR tetramer of the RPP1 resistosome. The catalytic site, BB-loop, AE interface and helices A, D, and E are labeled. The conformational change of BB-loop and ATP interaction with head-to-tail TIR dimer are highlighted. b, IDR prediction of TIR ___domain proteins by PONDR® server. c, AE interface mutations disrupt phase separation of TIR ___domain proteins. 1 mM NAD+ was incubated with 25 μM RPP1-TIRWT or RPP1-TIRS102D protein in the presence of 5% PEG 3,350 at 25 °C for 30 min. 0.2 mM NAD+ was incubated with 10 μM RBA1WT, RBA1AE, TX14WT, or TX14AE proteins at 25 C for 30 min. Scale bars = 20 μm. The experiments were repeated at least three times with similar results. d, Relative NADase activity of WT and AE mutant, RPP1-TIR, RBA1, and TX14 proteins. The NADase activity of WT TIR ___domain protein was normalized to 100%. Data are mean ± SD (n = 3 biologically independent samples). The concentrations of TIR proteins used for reactions were 40 μM. e, The input of the RPP1-TIR, RBA1, and TX14 variant proteins used for NADase assays. The experiments were repeated at least three times with similar results.
