Protein unfolding differential scanning

D. T. Haynie and E. Freire, Estimation of the folding/unfolding energetics of marginally stable proteins using differential scanning calorimetry, Anal. Biochem. 1994, 216, 33-41. 

Suggest a procedure for the measurement of the entropy of unfolding of a protein with differential scanning calorimetry (see In the laboratory 1.1). 

B. Chowdhry, S. Leharne. Simulation andAnalysis of Differential Scanning Calorimetry Output Protein Unfolding Studies 1. J. Chem. Educ. 1997, 74, 236-241. 

Simulation and Analysis of Differential Scanning Calorimetry Output Protein Unfolding Studies 1 87... 

The thermal unfolding of proteins is best measured by differential scanning calorimetry, which measures the heat absorbed by a protein as it is slowly heated through its melting transition (Figure 17.1). A solution of about 1 mg of protein in 1 mL of buffer and a separate reference sample of buffer alone are heated electrically.6 The additional current required to heat the protein solution is recorded. As the protein denatures, there is a large uptake of heat because the process is highly endothermic. The temperature at the maximum of the peak is... 

A study of two of the most prominent and widespread osmolytes, betaine and beta-hydroxyectoine, by differential scanning calorimetry (DSC) on bovine ribonu-clease A (RNase A) revealed an increase in the melting temperature Tm of RNase A of more than 12 K and of protein stability AG of 10.6 kj mol-1 at room temperature at a 3 M concentration of beta-hydroxyectoine. The heat capacity difference ACp between the folded and unfolded state was significantly increased. In contrast, betaine stabilized RNase A only at concentrations less than 3 M. When enzymes are applied in the presence of denaturants or at high temperature, beta-hydroxyectoine should be an efficient stabilizer. 

Fig. 4. Profile of a differential scanning calorimetry experiment done on a synthetic lysozyme. The heat capacity (kilocalories per degree per mole) of the unfolding process was monitored as a function of temperature on a Micro-Cal MC2 instrument. The transition midpoint of protein unfolding corresponds to the temperature at the peak of the curve, and the thermodynamic parameters A H and A Cp are evaluated by the procedure of Privalov.33...

Protein stability is the free energy difference (AG) between the folded and unfolded states at physiological conditions, and it is in the range of 5-25 kcal/mol. Site-directed mutagenesis experiments provided a wealth of data for understanding the importance of chemical interactions for the stability of proteins during amino acid substitutions. Protein stability is experimentally measured with differential scanning calorimetry, circular dichroism, fluorescence spectroscopy, and so forth. The availability of such data in an electronically accessible database would be a valuable resource for the analysis and prediction of protein mutant stability. 

The protein lysozyme, sm enzyme that brejiks down bactericd cell walls, unfolds at a tTcuisition temperature of 75.5 C, md the stcmdcffd enthalpy of transition as determined using differential scanning calorimetry is +509 kj mol". It follows that... 

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