Chain-like conformation of heat-denatured ribonuclease A and cytochrome c as evidenced by solution X-ray scattering.
BACKGROUND
Although the characterization of heat-denatured proteins is essential for understanding the thermodynamic mechanism of protein folding, their structural features are still unclear and controversial. In order to address this problem, we studied the size and shape of the heat-denatured states of bovine ribonuclease A (RNase A) and horse ferricytochrome c (cytochrome c) by solution X-ray scattering.


RESULTS
RNase A has four disulfide bonds, whereas cytochrome c, with a covalently bound heme group, has no disulfide bond. Guinier plots show that the heat-denatured RNase A is relatively compact, but the heat-denatured cytochrome c is expanded. On the other hand, the Kratky plots of the two proteins are similar, indicating that the heat-denatured proteins assume a chain-like disordered conformation. The X-ray scattering of RNase A and cytochrome c at various temperatures confirmed that their thermal transitions from a globular native state to a chain-like extended conformation can be approximated well by a two-state transition.


CONCLUSIONS
These results indicate that the heat-denatured RNase A and cytochrome c are substantially unfolded according to the criteria of solution X-ray scattering, although the heat-denatured RNase A remains compact because of the presence of the disulfide bonds. The results also confirm that the thermal denaturation occurs cooperatively with the breakdown of secondary and tertiary structure.
Hagihara
Y
Y
Department of Biology, Graduate School of Science, Osaka University, Toyonaka, Osaka 560, Japan.
Hoshino
M
M
Hamada
D
D
Kataoka
M
M
Goto
Y
Y
D000818:Animals; D002942:Circular Dichroism; D003574:Cytochrome c Group / Q000737:chemistry*; D006358:Hot Temperature; D008958:Models, Molecular; D010316:Particle Size; D011487:Protein Conformation; D011489:Protein Denaturation; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*; D012542:Scattering, Radiation; D012996:Solutions; D014965:X-Rays
D003574:Cytochrome c Group; D012996:Solutions; D012259:Ribonuclease, Pancreatic