Tesis doctoral de Josep Font Sadurní
Using temperature and pressure as denaturants we have explored the contributions to stability of the hydrophobic core residues of rnase a with the positions most critical for stability being v54, v57, i106 and v108. It is also shown that the stability differences can be attributed to both hydrophobic interactions and packing density with an equivalent energetic magnitude. These results are consistent with an exquisite tight core packing and the existence of rearrangements in the protein interior even after drastic deleterious substitutions. the role of hydrophobic interactions established by the residues that belong to the main hydrophobic core of rnase a in its pressure-folding transition state, was investigated using the phi-value method. The folding kinetics was studied using pressure-jump techniques both in the pressurization and depressurization directions. The structure of the transition state of the hydrophobic core of rnase a, from the point of view of formed interactions, is a relatively, uniformly expanded form of the folded structure with a mean phi f -value of 0.43. This places it half way between the folded and unfolded states. On the other hand, for the variants, the average of beta p-values is 0.4, suggesting a transition state that is 40% native-like. Altogether the results suggest that the pressure-folding transition state of rnase a, looks like a collapsed globule with some secondary structure and a weakened hydrophobic core. pressure-jump induced relaxation kinetics was used to explore the energy landscape of protein folding/unfolding of y115w variant of rnase a. Whereas downward p-jumps resulted always in single exponential kinetics, the kinetics induced by upward p-jumps was biphasic in the low pressure range (below p1/2) and monophasic at higher pressures. At 50 °c, only one phase was observable. Analysis of the activation volume of the fast phase suggests a temperature dependent structural shift of the unfolding transition state. The lat
Datos académicos de la tesis doctoral «Role of the main hydrophobic core of rnase a on the folding and unfolding processes induced by pressure and temperature.«
- Título de la tesis: Role of the main hydrophobic core of rnase a on the folding and unfolding processes induced by pressure and temperature.
- Autor: Josep Font Sadurní
- Universidad: Girona
- Fecha de lectura de la tesis: 02/06/2006
Dirección y tribunal
- Director de la tesis
- María Vilanova Brugués
- Tribunal
- Presidente del tribunal: claudi Cuchillo foix
- krzysztof Lewinski (vocal)
- f. Lange hans (vocal)
- v. Laurents douglas (vocal)