Salt-induced formation of the molten globule state of cytochrome c studied by isothermal titration calorimetry.


Abstract

Although the molten globule state has been proposed as a major intermediate of protein folding, it has proven difficult to obtain thermodynamic data characterizing this state. To explore another approach for characterizing the molten globule state, salt-induced formation of the molten globule state of horse cytochrome c at pH 1.8 was studied by isothermal titration calorimetry. By titrating the acid-unfolded cytochrome c with sodium perchlorate, an exothermic reaction was observed. The titration curve obtained from the heat was cooperative and agreed well with the conformational transition curve measured by CD at 222 nm. This result indicated that the salt-induced conformation change is well approximated by a two-state transition between the acid-unfolded and molten globule states. The heat for formation of the molten globule state estimated by isothermal titration calorimetry was consistent with the enthalpy change for unfolding of the sodium perchlorate-stabilized molten globule state at pH 1.8, which was measured by differential scanning calorimetry and CD. These results indicate that the heat of titration largely reflects the enthalpy change of the conformational transition. From these results, we consider that isothermal titration calorimetry will become a useful approach for investigating the molten globule state. Study holds ProTherm entries: 10958, 10959, 10960, 10961, 10962, 10963, 10964, 10965, 10966 Extra Details: compact denatured state; anion binding; protein folding;,thermal unfolding

Submission Details

ID: HaaKTwL93

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:41 p.m.

Version: 1

Publication Details
Hamada D;Kidokoro S;Fukada H;Takahashi K;Goto Y,Proc. Natl. Acad. Sci. U.S.A. (1994) Salt-induced formation of the molten globule state of cytochrome c studied by isothermal titration calorimetry. PMID:7937949
Additional Information

Number of data points 32
Proteins Cytochrome c ; Cytochrome c
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: dCp ionic:NaClO4: 100 mM, prot_conc:0.1 mg/ml ; Experimental Assay: Tm ionic:NaClO4: 100 mM, prot_conc:0.1 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 100 mM, prot_conc:0.1 mg/ml ; Experimental Assay: dCp ionic:NaClO4: 50 mM, prot_conc:0.1 mg/ml ; Experimental Assay: Tm ionic:NaClO4: 50 mM, prot_conc:0.1 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 50 mM, prot_conc:0.1 mg/ml ; Experimental Assay: dCp prot_conc:0.1 mg/ml, ionic:NaClO4: 20 mM ; Experimental Assay: Tm prot_conc:0.1 mg/ml, ionic:NaClO4: 20 mM ; Experimental Assay: dHvH prot_conc:0.1 mg/ml, ionic:NaClO4: 20 mM ; Experimental Assay: dCp prot_conc:0.1 mg/ml, ionic:NaClO4: 10 mM ; Experimental Assay: Tm prot_conc:0.1 mg/ml, ionic:NaClO4: 10 mM ; Experimental Assay: dHvH prot_conc:0.1 mg/ml, ionic:NaClO4: 10 mM ; Experimental Assay: dCp ionic:NaClO4: 60 mM, prot_conc:5 mg/ml ; Experimental Assay: dHcal ionic:NaClO4: 60 mM ; Experimental Assay: Tm ionic:NaClO4: 60 mM, prot_conc:5 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 60 mM, prot_conc:5 mg/ml ; Experimental Assay: dCp ionic:NaClO4: 50 mM, prot_conc:5 mg/ml ; Experimental Assay: dHcal ionic:NaClO4: 50 mM ; Experimental Assay: Tm ionic:NaClO4: 50 mM, prot_conc:5 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 50 mM, prot_conc:5 mg/ml ; Experimental Assay: dCp ionic:NaClO4: 40 mM, prot_conc:5 mg/ml ; Experimental Assay: dHcal ionic:NaClO4: 40 mM ; Experimental Assay: Tm ionic:NaClO4: 40 mM, prot_conc:5 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 40 mM, prot_conc:5 mg/ml ; Experimental Assay: dCp ionic:NaClO4: 30 mM, prot_conc:5 mg/ml ; Experimental Assay: dHcal ionic:NaClO4: 30 mM ; Experimental Assay: Tm ionic:NaClO4: 30 mM, prot_conc:5 mg/ml ; Experimental Assay: dHvH ionic:NaClO4: 30 mM, prot_conc:5 mg/ml ; Experimental Assay: dCp prot_conc:5 mg/ml, ionic:NaClO4: 20 mM ; Experimental Assay: dHcal ionic:NaClO4: 20 mM ; Experimental Assay: Tm prot_conc:5 mg/ml, ionic:NaClO4: 20 mM ; Experimental Assay: dHvH prot_conc:5 mg/ml, ionic:NaClO4: 20 mM
Libraries Mutations for sequence GDVEKGKKIFVQKCAQCHTVEKGGKHKTGPNLHGLFGRKTGQAPGFTYTDANKNKGITWKEETLMEYLENPKKYIPGTKMIFAGIKKKTEREDLIAYLKKATNE
Sequence Assay Result Units