FIGURE 4.

¹⁵N HSQC NMR spectra of E_CT1 and E_CT1H_CT. Two-dimensional ¹H/¹⁵N HSQC NMR spectra taken at 298 K in 25 mm sodium phosphate (pH 7), 0.5 mm EDTA, and 0.02% NaN₃. Spectra presented are: E_CT1 (A), E_CT1H_CT where H_CT is unlabeled for clarity (B), and where E_CT1H_CT is fully labeled (red) and is overlaid by the partially labeled spectrum from B in blue (C). The observed change in E_CT1 behavior upon H_CT binding, explored in Fig. 3, is further evidenced here by the global improvement in backbone amide peak signal dispersion and homogeneity indicative of E_CT1 fold stabilization by H_CT binding. Side chain amide peaks are linked by black horizontal lines. Resonances marked with an asterisk belong to amino acid side chains (e.g. Arg) or residues that could not be assigned because of overlap/poor signal to noise. KLi and VLi are part of the N-terminal linker (GPKVP). For E_CT1 and E_CT1H_CT, Ser¹¹³, Gly¹⁴⁸, Ile¹⁶⁴, Ser¹³⁴, Asp¹³⁵, and Gly¹³⁶ could not be assigned, respectively. The mutation Ta_E C121A has been described previously (Ref. 12; indicated by the red label in B). The concentration of NMR samples was between 0.5 and 1 mm.