Fig. 1
From: Dynamic structural states of ClpB involved in its disaggregation function
Diverse oligomeric forms of ΔN-TClpB observed by HS-AFM. a HS-AFM image acquired in 0.5 mM ATP at 25 °C. Scale bar, 20 nm. Z color bar, 0 to 9.2 nm. b–e Four typical oligomeric forms observed in the presence of ATP: b round closed ring, c spiral, d twisted-half-spiral, and e open ring. We collectively refer to the spiral and twisted-half-spiral rings as the distorted closed ring. Scale bar, 5 nm. Z color bar, 0 to 8.0 nm. f Cryo-EM structure of spiral hexamer of Hsp104 without N-terminal domains (PDB code: 5KNE)39 and schematic of the spiral architecture. g Simulated AFM image of N-terminal-truncated spiral hexamer of Hsp104 constructed using a cone-shaped probe with a tip radius of 0.5 nm and a half cone angle of 5°. The constructed image was filtered by a low-pass filter with a cut-off frequency of 3 nm. Scale bar, 5 nm. Z color bar, 0 to 9.3 nm. h Histograms of circularity for the oligomers formed at different [ATP]. Numbers of molecules analyzed are 425 (10 μM), 389 (50 μM), 438 (0.1 mM), 415 (0.5 mM), 624 (1 mM), and 410 (2 mM). i Percentages of closed (black area) and open (hatched area) forms at various [ATP] estimated from the histograms. A threshold value of circularity was set at 0.68. Percentages of the closed and open forms observed at 1 mM ATPγS and 1 mM ADP are also displayed (413 molecules for ATPγS and 432 molecules for ADP). j Percentage of closed rings as a function of [ATP] (open circles). The data points were fitted by a simple 1:1 binding model with an apparent Kd = 9.9 ± 1.8 μM (mean ± s.d.) (black line). The magenta line shows ATP occupancy in the ATP-binding sites of ΔN-TClpB hexamers theoretically calculated using an apparent Kd (9.2 μM) estimated by replacement titration of bound mant-ADP with ATP (Supplementary Figure 8)
