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. Author manuscript; available in PMC: 2013 Nov 1.
Published in final edited form as: Biochim Biophys Acta. 2012 Apr 16;1823(11):2046–2056. doi: 10.1016/j.bbamcr.2012.04.003

Figure 4.

Figure 4

NMR studies of the interaction between Ub and hHR23a UBA2 at pH 4.5. (A) Chemical shift perturbations in the individual amides in Ub caused by UBA2 binding. (B) Chemical shift perturbations in the individual amides in UBA2 caused by Ub binding. Data in both (A) and (B) correspond to the endpoints in titration. The horizontal bars on top of the plot in (B) indicate the location of UBA2’s α-helices. (C) Overlay of 1H-15N HSQC spectra of 15N-labeled UBA2 at several titration points with unlabeled Ub; shown are regions corresponding to signals of G331 and S335. (D) Representative titration curves (symbols) and their fit to a 1:1 binding model (curves) for selected amides in 15N-labeled UBA2 upon addition of Ub. The Kd value averaged over 11 residues was 215 ±35 μM. (E, F) Map of the perturbations in Ub and UBA2 observed in this study (pH 4.5) on the structure of the Ub/UBA2 complex obtained at neutral conditions [25, 50]. UBA2 is shown as a ribbon (cyan), while Ub is colored green and shown as a ribbon in E and as a surface in F; the perturbed residues are colored red (Ub) and blue (UBA2).