Figure 2. Impact of nucleotides on the conformational dynamics of SecA.
(a) Structures of the complex highlighting the contact sites (cyan) in SecA with SecG (green), SecY (pink), mature regions of the pre-protein (PP-M; grey) and ATP (orange). Significant sum differences in relative deuterium uptake (ΔHDX = AMPPNP-ADP) of (b) SecA without SecYEG, and (c) of SecA in the presence of a molar excess of SecYEG after 30 min of deuteration. Highlighted regions represent contacts with SecG, SecY, pre-protein (mature domain, PP-M) and ATP; coloured according to (a). Dashed boxes in (c) highlight regions in contact with the pre-protein mature domain (PP-M). Detailed information of HDX-MS data is provided in Supplementary files 1a and 1b.
Figure 2—figure supplement 1. Linear protein coverage maps for SecA, SecY, SecE and SecG.
Maps were generated using Deuteros (Lau et al., 2019).
Figure 2—figure supplement 2. Woods plots illustrating the impact of nucleotides on the conformational dynamics of SecA.
Woods plots comparing the relative deuterium uptake (AMPPNP-ADP) of (a) SecA alone and (b) SecA in complex (and saturated) with SecYEG after incubation at 25°C in deuterated solvent for 15 s, 1, or 5 min. The dotted lines represent the 99% confidence interval, which indicates that the level of difference of deuterium uptake between the two compared states is statistically significant. Bars represent individual peptides. Bar length corresponds to peptide size. Red and blue coloured bars indicate statistically significant deprotected or protected peptides, respectively.
Figure 2—figure supplement 3. Impact of SecYEG binding on SecA.
Woods plots comparing the relative deuterium uptake of SecA upon interaction with SecYEG (present in excess) at 15 s, 1, 5 or 30 min. Results indicate an overall deprotection, hence destabilisation of SecA upon SecYEG binding.