Figure 5
From: Conformational rearrangements of the C1 ring in KaiC measure the timing of assembly with KaiB
Fluorescence detection of the C1-ring rearrangements in the presence of KaiB. (a) Apparent fluorescence intensity (FIapp) of the phospho-mimicking mutants, S/E, D/E, D/T and A/A of KaiCS157W after 24 h incubation at 30 °C with or without an equimolar concentration of KaiB (monomer basis). Each dataset is shown in a different color as a pair of filled circles connected with a line, and the mean value from independent measurements is represented as bars. (b) Gel-filtration chromatograms of mixtures containing KaiB and KaiCS157W-A/A (green) or KaiCS157W-D/E (blue). The samples were loaded onto a column immediately after the mixing (dashed lines, t = 2 min) or after 18 h incubation at 30 °C (solid lines, t = 18 h). Each trace was vertically shifted for clarity of presentation. (c) SDS-PAGE analysis of peaks a–g in (b). Raw image is shown in Supplementary Fig S6. (d) Time course of FIapp for KaiCS157W-D/E before and after addition of an equimolar concentration of KaiB. The displayed trace corresponds to a green-colored dataset of KaiCS157W-DE in (a). A time of addition of KaiB was set as t = 0 h.
