Partially unfolded equilibrium state of hen lysozyme studied by circular dichroism spectroscopy.
Equilibrium unfolding of hen egg white lysozyme as a function of GdnCl concentration at pH 0.9 was studied over a temperature range 268.2-303.2 K by means of CD spectroscopy. As monitored by far- and near-UV CD at 222 and 289 nm, the lack of coincidence between two unfolding transition curves was observed, which suggests the existence of a third conformational species in addition to native and unfolded states. The three-state model, in which a stable intermediate is populated, was employed to estimate the thermodynamic parameters for the GdnCl-induced unfolding. It was found that the transition from the native to intermediate states proceeds with significant changes in enthalpy and entropy due to an extremely cooperative process, while the transition from the intermediate to unfolded states shows a low cooperativity with small enthalpy and entropy changes. These results indicate that the highest energy barrier for the GdnCl-induced unfolding of hen lysozyme is located in the process from the native state to the intermediate state, and this process is largely responsible for the cooperativity of protein unfolding.
Sasahara
K
K
Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Japan.
Demura
M
M
Nitta
K
K
D000818:Animals; D002645:Chickens; D002942:Circular Dichroism; D005260:Female; D019791:Guanidine / Q000494:pharmacology; D007700:Kinetics; D008956:Models, Chemical; D009113:Muramidase / Q000737:chemistry* / Q000187:drug effects; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
D009113:Muramidase; D019791:Guanidine