Thermodynamic characterization of the folding process of Psychromonas ingrahamii?s frataxin (pFXN) by using calorimetry and circular dichroism

BATASTINI, N.A.; FL. GONZÁLEZ FLECHA; GONZÁLEZ LEBRERO, RODOLFO M.; EA. ROMAN

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofísica.; 2018

Sociedad Argentina de Biofísica.

Despite proteins in a family share certain degree of sequence identity andstructural conservation, the process by which each one reaches the nativestructure may be specifically and subtle regulated. In this sense, frataxin familyshowing a general structural conservation but different celular roles betweeneukaryotes and prokaryotes, display several folding mechanism under the sameconditions. Frataxins are around 7-10 kcal/mol stable but depending on themolecular species present, particular signatures in its stability modulation wereobserved. Psychromonas ingrahamii?s frataxin (pFXN) shows a pH dependence ofthe folding Gibbs free energy when going from pH 6 to 8.The main goal of this work is to perform a thermodynamic characterization of thepFXN folding process and its pH and temperature dependencies. Here, usingisothermal titration calorimetry (ITC) we measure the exchanged heat during pFXNfolding as a function of pH and temperature, and parallely, secondary structurechanges were registered by far-UV ellipticity measurements.In a first approach, the change in the measured heat was analyzed in terms ofthree components: the protonation of the native and denatured states, and thefolding of the protein. The enthalpy change for this last process (dHf) was modeledas a first order Taylor series of pH. Circular dichroism experiments yielded thenative and denatured fractions of pFXN as a function of pH at differenttemperatures and the number of protons involved in the process. This analysisalso allowed the calculation of the dependency of the folding entropy on pH. Ourresults points to an exhaustive characterization of the folding process of pFXN witha robust thermodynamic description of the process yielding not only the foldingenthalpy and entropy but also their derivatives respect to the pH and the changesin heat capacity for the folding process and for basal states.AcknowldegmentsPICT-2016, Ernesto A. Roman, Agencia Nacional de Promoción Científica y Tecnológica.