Figure 9: Ca²⁺ induced the solvent exposure of the signalling tyrosine in the ITT motif.

(a) Superimposed ¹⁵N–¹H TROSY spectra of mIgG-tail with POPG bicelles, mIgG-tail with POPG bicelles and different concentrations of Ca²⁺, and mIgG-tail in solution as a control. Membrane-sequestered mIgG-tail and solvent-exposed mIgG-tail had very different amide resonance spectra. Ca²⁺ was titrated into the mIgG-tail with POPG sample at a molar ratio of [Ca²⁺]: [POPG] from 0 to 0.5. In response to the increase in the Ca²⁺ concentration, the mIgG-tail amide resonances exhibited systematic shifts from the membrane-bound state to the solvent-exposed state. G26 is shown as an example of Ca²⁺ titration effects (upper dotted box). (b) Strips from aromatic NOESY spectra showing NOEs (distance of <5 Å) between the aromatic protons of W5, F7 and Y21 and the methylene protons in the lipid acyl chains. The substantial intermolecular NOE signals (marked by asterisks) observed in the absence of Ca²⁺ indicated the insertion of tryptophan, phenylalanine and tyrosine side chains into the membrane hydrophobic interior. The addition of Ca²⁺ resulted in the significant decrease of NOE signals, which indicated that side chains of these three residues were dissociated from the membrane core region.