FIGURE 6.

Analysis of E_CT1 chemical shift change upon the addition of H_CT and a model of the proposed domain arrangement in the full-length EH peripheral stalk complex. Assignment of the ¹⁵N HSQC spectra presented in Fig. 4 permitted the calculation of E_CT1 amide-proton chemical shift changes (Δ¹Hδ) upon binding H_CT. A, as can be seen, the majority of residues experiencing the most significant chemical shift perturbation upon H_CT binding are located in the N- and C-terminal helices of E_CT1 (residues for which no assignments could be obtained in either the E_CT1 (Ser¹¹³, Gly¹⁴⁸, and Ile¹⁶⁴) or E_CT1H_CT (Ser¹³⁴, Asp¹³⁵, and Gly¹³⁶) spectra are marked with an asterisk). B, the positions analogous to the Ta_E_CT1 residues undergoing the largest chemical shift changes are highlighted in the available crystal structure of Ph_E_CT with one dimer partner removed (Protein Data Bank code 2dma; Ref. 11). The largest change in chemical shift in the middle portion of the structure is experienced by Tyr¹⁴⁶, which can be seen located close to the N- and C-terminal α-helices of Ph_E_CT in the crystal structure (Ta_E Y146 corresponds to Ph_E M157). C, H_CT, modeled as an α-helix and guided by the data presented here, is docked onto the Ph_E_CT crystal structure monomer. Continuing below the globular E_CT1H_CT domain, the N-terminal domain, E_NT2H_NT, is modeled as a pair of parallel helices, representing the coiled-coil domain. We speculate that the N-terminal α-helix of H_NT is folded back to interact with and stabilize the N-terminal ends of the coiled-coil domain. D, the resulting EH model placed into a schematic model of the T. acidophilum A-ATPase. The stoichiometry of the EH peripheral stalks has recently been determined for the related A/V-ATPase from T. thermophilus (6).