Native exchange NMR None ΔGu =~9.6 kcal/mol ADLEDNWETLNDNLKVIEKADNAAQVKDALTKMRAAALDAQKATPPKLEDKSPDSPEMKDFRHGFDILVGQIDDALKLANEGKVKEAQAAAEQLKTTIRAYNQKYG

The exchange reaction was initiated by dissolving protein samples with deuterated amide protons in an exchanging solution using a spin column filled with G25 Sephadex and preequilibrated with exchanging buffer (50 mM NaAc-d, 95% H2O, and 5% D2O). The deuterated Rd-apocyt b562 protein was prepared by dissolving proteins in D2O (pD 7.0, 0.1 M Na2PO4) and heated at 60 °C for more than 30 min. 15N-1H HSQC spectra were recorded using a 500 MHz instrument. Protein concentrations are 2 mM. The peak intensity as a function of time was measured and fitted to single-exponential kinetics to obtain the exchange rate constant.

Native exchange NMR None ΔGu =~6.5 kcal/mol ADLEDNWETLNDNLKVIEKADNAAQVKDALTKMRAAALDAQKATPPKLEDKSPDSPEMKDFRHGFDILVGQIDDALKLANEGKVKEAQAAAEQLKTTIRAYNQKYG

The exchange reaction was initiated by dissolving protein samples with deuterated amide protons in an exchanging solution using a spin column filled with G25 Sephadex and preequilibrated with exchanging buffer (50 mM NaAc-d, 95% H2O, and 5% D2O). The deuterated Rd-apocyt b562 protein was prepared by dissolving proteins in D2O (pD 7.0, 0.1 M Na2PO4) and heated at 60 °C for more than 30 min. 15N-1H HSQC spectra were recorded using a 500 MHz instrument. Protein concentrations are 2 mM. The peak intensity as a function of time was measured and fitted to single-exponential kinetics to obtain the exchange rate constant.

Native exchange NMR None ΔGu =~4.5 kcal/mol ADLEDNWETLNDNLKVIEKADNAAQVKDALTKMRAAALDAQKATPPKLEDKSPDSPEMKDFRHGFDILVGQIDDALKLANEGKVKEAQAAAEQLKTTIRAYNQKYG

The exchange reaction was initiated by dissolving protein samples with deuterated amide protons in an exchanging solution using a spin column filled with G25 Sephadex and preequilibrated with exchanging buffer (50 mM NaAc-d, 95% H2O, and 5% D2O). The deuterated Rd-apocyt b562 protein was prepared by dissolving proteins in D2O (pD 7.0, 0.1 M Na2PO4) and heated at 60 °C for more than 30 min. 15N-1H HSQC spectra were recorded using a 500 MHz instrument. Protein concentrations are 2 mM. The peak intensity as a function of time was measured and fitted to single-exponential kinetics to obtain the exchange rate constant.