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

Thermodynamic insights into the binding of ANS with the salt induced molten globule states of cytochrome c

“…The increase in the secondary structural content of cyt c from the acid unfolded state with rising concentration of NaClO 4 in solution has been reported by Hamada et al [23]. They have observed complete induction of the MG-state in the protein in presence of 0.1 mol Á dm À3 salt.…”

“…Cyt c, like other acid denatured proteins is maximally unfolded at pH 2.0 by HCl in the absence of salts and undergoes a cooperative transformation to an intermediate structure called the A-state [18] which has properties typical of the molten globule [5,19] upon addition of anions from either salts or acids. The molten globule state or the A-state of cyt c has been studied extensively in recent years by a variety of techniques which has enabled its main features to be defined [15,16,[20][21][22][23]. The A-state of cyt c has been reported in acidic solution containing high salt concentration [2,14,24] or through neutralization of charges by acetylation [25,26].…”

Structural characterization of the molten globule of α‐lactalbumin by solution X‐ray scattering

“…The thermal unfolding of the molten globule state of cytochrome c and apomyoglobin involves a cooperative transition with an enthalpy change and heat capacity change of unfolding (Potekhin & Pfeil, 1989;Kuroda et al, 1992;Hagihara et al, 1994;Hamada et al, 1994;Nishii et al, 1994Nishii et al, , 1995. Cooperative thermal unfolding with distinct heat absorption has also been reported for the staphylococcal nuclease fragment (Gittis et al, 1993;Griko et al, 1994a).…”

Unfolding of Rabbit Muscle Creatine Kinase Induced by Acid

“…The positive molar heat capacity change associated with the unfolding of a protein, for example, is commonly attributed to the hydrophobic interaction, although other factors may contribute to ⌬ U C P,m (41,57). A large positive molar heat capacity change of the unfolding of MM-CK induced by acid, 8.78 kcal mol Ϫ1 K Ϫ1 , at all temperatures examined indicates that hydrophobic interaction is the dominant driving force stabilizing the native structure of this protein.…”