Thermodynamic characterisation of RNAase a in the presence of urea and GuHCl |
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Authors: | G. Barone P. Del Vecchio D. Fessas C. Giancola G. Graziano A. Riccio |
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Affiliation: | (1) Department of Chemistry-University ‘Federico II’, Via Mezzocannone 4, 80134 Naples, Italy |
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Abstract: | It is presented a study concerning the influence of guanidinium chloride (GuHCl) and urea on thermal stability of Bovine Pancreatic Ribonuclease A (RNAase A) at differentpH values. As expected, at increasing the denaturant concentration, the protein thermostability decreases. This is shown by a decrease of both the thermodynamic parameters, temperature and heat effect, characterising the denaturation process. In order to analyse the calorimetric curves we adopt a statistical thermodynamic approach. The individual one-dimensional DSC profiles have been expanded into another dimension by varying the GuHCl concentration, so that a heat capacity surface is defined for eachpH. By means of the ICARUS program, developed in our laboratory, we accomplish a two dimensional deconvolution of the experimental data linking the binding equilibrium to the denaturation process. This analysis provides a well founded and complete statistical thermodynamic characterisation of denaturation process of RNAase A in the presence of GuHCl and allows to calculate the thermodynamic parameters associated to the binding of denaturant molecule. |
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Keywords: | denaturation models guanidinium chloride micro-DSC ribonuclease A urea |
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