NMR analyses of ALIX-CHMP4 interactions and CHMP4 conformtation and dynamics

Figure 4A: 1H/15N chemical shift perturbation profiles of 100 μM 15N/2H-labeled Bro1 on addition of 300 μM unlabeled CHMP4 constructs; CHMP4C(121–233, S191C) and CHMP4B(121–224,S184C). Figure 4C: 1H/15N and 13C’/15N chemical shift perturbation profiles of 150 μM 15N/13C-labeled PRD(703–800)Strep in the presence of 450 μM unlabeled CHMP4 constructs. Figure 5F: Evidence of transient long-range interactions in CHMP4C paralog using experimental PRE profiles (shown as paramagnetic-labeled vs. diamagnetic-labeled 1H/15N cross-peak height ratios) of 15N-labeled CHMP4C(121–233,G154C) and CHMP4C(121–233,M165C). Figure 6A and 6B: The reduction in 1H/15N cross-peak heights of 150 μM 15N-labeled (A) CHMP4C(121–233,S191C) and (B) CHMP4B(121–224,S184C) as a function of increasing amount of unlabeled Bro1. Figure 6C and 6D: The perturbations in 1H/15N chemical shifts of 150 μM 15N-labeled (C) CHMP4C(121–233,S191C) and (D) CHMP4B(121–224,S184C) as a function of increasing amount of unlabeled PRD(703–800)Strep. Figure 6I, 6J, and 6K: The reduction in 1H/15N cross-peak heights of 30 μM 15N/2H-labeled (I) (CHMP4C121–233,S191C), (J) CHMP4B(121–224,S184C), and (K) CHMP4C(121–224,S184C, I225A, L228A, W231A) in the presence of 90 μM unlabeled ALIX(1–868,P801G)Strep.