Denaturation of a recombinant cutinase from Fusarium solani in AOT-iso-octane reverse micelles: A steady-state fluorescence study

Near UV absorbance and fluorescence spectroscopy show conformational changes of a recombinant cutinase from Fusarium solani incorporated in sodium-di-2-ethylhexyl sulfosuccinate (AOT)-iso-octane reversed micelles with W-0 = [H2O]/[AOT] = 20. Excitation spectra were used to decompose cutinase absorbance in its Trp and Tyr components, showing that the latter absorb red-shifted in the native cutinase in aqueous solution as compared to free Tyr, whereas in reverse micelles and denatured cutinase no shift is detected. Emission maxima variations (lambda(max) 303, 311 and 335 nm, respectively in aqueous, reverse micelles and thermally denatured cutinase) reflect progressive changes in the micropolarity of the environment and exposure of Trp residues at the protein surface. The encapsulation of cutinase in AOT-iso-octane reversed micelles induces a time-dependent denaturation measured by fluorescence intensity changes at 330 nm, which match the profile of enzyme activity loss in this media.