The structural stability of cyclophilin A （CypA） was investigated using H/D exchange and temperature coefficients of chemical shifts of amide protons, monitored by 213 heteronuclear NMR spectroscopy. Amide proton exchange rates were measured by H/D exchange experiments for slow-exchange protons and measured by SEA （Solvent Exposed Amides）-HSQC experiments for fast-exchange protons. Temperature coefficients of chemical shifts and hydrogen exchange rates of amide protons show reasonably good correlation with the protein structure. Totally, 44 out of 153 non-proline assigned residues still exist in 86 d of hydrogen-deuterium exchange at 4 ℃, suggesting that CypA structure should be highly stable. Residues in secondary structures of α2, β1, β2, β5, β6 and β7 might constitute the hydrophobic core of the protein. The change in free energy of unfolding （ △Gu^H2O ） of CypA was estimated to be （21.99± 1.53） kJ·mol^-1 by circular dichroism （CD）. The large free energy change is also an indicator of the high structural stability.
