Global and local structural changes of cytochrome c and lysozyme characterized by a multi-group unfolding process

¹ Institute of Atomic and Molecular Sciences, Academia Sinica, Taiwan
² NSRRC
³ National Synchrotron Radiation Research Center, Taiwan
⁴ National Synchrotron Radiation Research Center, Taiwan
⁵ Institute of Chemistry, Academia Sinica, Taiwan
⁶ Department of Chemistry, National Taiwan University, Taiwan
⁷ Department of Chemical Engineering, National Tsing Hua University, Taiwan

^* To whom correspondence should be addressed. E-mail: usjeng{at}nsrrc.org.tw.

Submitted on October 22, 2007
Revised on November 17, 2007
Accepted on 30 January 2008

	Abstract

Equilibrium unfolding behaviors of cytochrome c and lysozyme induced by presence of urea (0 to 10 M) as well as changes in temperature (295 to 363 K) or pH (1.8 to 7) are examined via small-angle X-ray scattering (SAXS) and spectroscopic techniques including circular dichroism (CD) and optical absorption. Denaturant and temperature effects are incorporated in the free energy expression for a general multi-group unfolding process. Results indicate that there are at least four unfolding groups in temperature-, urea- or pH-induced unfolding of cytochrome c: two of these are related to the prosthetic heme group whereas the other two correspond respectively to unfolding of -helices and global changes in protein morphology that are largely unaccounted for by the first two groups. In contrast, unfolding of lysozyme approximately follows a simple one-group process. A modified mean-field Ising model is adopted for a coherent description of the unfolding behaviors observed. Thermodynamic parameters extracted from simple denaturing processes, on the basis of the Ising model, can closely predict unfolding behaviors of the proteins in compounded denaturing environments.

Key Words: Ising model, Multi-group unfolding, SAXS, cytochrome c, lysozyme