Figure 6.

Dynamic versus Rigid Body Interactions in Different Protein Complexes

(A) The ratio of the H/D exchange rates of ¹⁵N-labeled mβ₂m bound (∼22%) to unlabeled ΔΝ6 (80 μΜ mβ₂m + 40 μΜ ΔΝ6) versus free (80 μΜ) ¹⁵N-labeled mβ₂m (k_ex ratio, bound:free) plotted against residue number. An increase in k_ex ratio indicates a loss of H/D exchange protection upon binding. Error bars represent the propagated error of the fits to the raw data shown in Figure S6.

(B) As in (A) but for free (80 μΜ) ¹⁵N-labeled hβ₂m versus ∼22% ¹⁵N-labeled hβ₂m bound to ΔΝ6 (80 μΜ hβ₂m + 160 μΜ ΔΝ6). Note that exchange of hβ₂m occurs by a mixed EX1/EX2 mechanism (Hodkinson et al., 2009), ruling out analysis of these data in terms of the free energy of binding.

(C) Differences in ¹H (gray) and ¹⁵N (red) chemical shifts when ¹⁵N-labeled ΔΝ6 and ¹⁴N-labeled mβ₂m are mixed in a 1:1 ratio (80 μΜ each; ∼45% ΔΝ6-bound). Dotted lines represent two standard deviations of the mean over the entire data set for each nucleus.

(D) As in (C) but for ¹⁵N-labeled ΔΝ6 mixed with ¹⁴N-labeled hβ₂m (80 μΜ hβ₂m + 480 μΜ ΔΝ6; ∼45% ΔΝ6-bound). Black bars denote residues that are broadened beyond detection but show significant chemical shift changes when less mβ₂m/hβ₂m is added. Red crosses denote ΔΝ6 residues that are broadened at pH 6.2. Residues that experience significant chemical shift changes on binding are highlighted in blue and pink backgrounds in (C) and (D), respectively.