Solvent tunes the peroxidase activity of cytochrome c immobilized on kaolinite

• Cytochrome c strongly adsorbs on kaolinite in presence of ethanol.
• Two adsorbed forms are identified: a molten globule and a helical state.
• The peroxidase activity of adsorbed cytochrome c depends on ethanol concentration.
• Adsorption conditions affect the peroxidase activity of the protein.
• The molten globule form is found to be the most catalytically active.

The adsorption process and the peroxidase activity of yeast cytochrome c (ycc) immobilized on kaolinite (Kaol) were investigated in mixed ethanol/water solutions. The protein strongly adsorbs on the surface of the clay mineral and the thermodynamic adsorption constant increases with increasing ethanol concentration. The adsorption parameters suggest that in ycc a conformational transition from molten globule to helical state occurs in solution for ethanol concentration above 20%. The peroxidase activity of ycc immobilized on Kaol increases from 0% to 20% ethanol (v/v), then it progressively decreases and almost vanishes in pure ethanol. The catalytic properties of adsorbed ycc were studied in 20 and 40% ethanol solutions which correspond to the molten globule and to the helical state, respectively. In both cases, catalysis adheres to the Michaelis–Menten model. The molten globule state, which binds more weakly to kaolinite than the helical state, was found to be more catalytically active. This study is meant to identify the physicochemical factors that modulate the catalytic activity of this kaolinite-based interface of broad applicability.

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