IQUIFIB   02644
INSTITUTO DE QUIMICA Y FISICOQUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Protonation effects on folding of a psychrophilic frataxin variant
Autor/es:
ERNESTO A. ROMAN; LUCAS DEFELIPE
Lugar:
Capital federal
Reunión:
Congreso; International Society for Computational Biology meeting; 2016
Institución organizadora:
International Society for Computational Biology - A2B2C
Resumen:
Here, we will study the stabilization mechanisms and dynamics of frataxin scaffold using a variant from Pyschromonas ingrahamii as a model (pFXN). We previously reported that pFXN stability is highly modulated by pH in the range 6-8 (Roman EA at al. Biochim. Biophys. Acta, 2013). Here, we will discuss our results on the thermodynamic and kinetic stability of pFXN by rapid mixing experiments. Folding and unfolding dependence on pH behave differently leading to two apparent pKa for native and unfolded states. Models with single protonation sites for the native state were discarded. This implies that at least different group of residues or different conformations exists in the native state. Apparent pKa for native and unfolded states around 8.4 and 5.0, respectively. This suggests that a strong shift in the pKa of a residue/s upon folding is associated with the pH stabilization. In addition, temperature equilibrium chemical unfolding experiments at different pHs showed a bell-shaped dependence of the Gibbs Free Energy between 4 and 26 Celsius degrees. The thermodynamic analysis suggests that the increase in stability comes mainly from a change in the entropy energy term. This may be explained by residual structure in the unfolded state, local unfolding in the native state, but also from difference in the solvation shells due to charge states in both ensembles. Constant pH simulations showed that besides histidine residues there are some glutamic acid residues located in the main loop that have their pKa shifted towards pH 6-7. This result is in agreement with the fact that His point mutants are not able to revert the stability dependence on pH. Preliminary NMR results show that a complex network of hydrogen bonded interactions are strongly regulated by pH in in the range between 6 and 8