* To whom correspondence should be addressed.
Received March 25, 2009; Revision received April 20, 2009
Equilibrium unfolding of stem bromelain (SB) with urea as a denaturant has been monitored as a function of pH using circular dichroism and fluorescence emission spectroscopy. Urea-induced denaturation studies at pH 4.5 showed that SB unfolds through a two-state mechanism and yields ΔG (free energy difference between the fully folded and unfolded forms) of ~5.0 kcal/mol and Cm (midpoint of the unfolding transition) of ~6.5 M at 25°C. Very high concentration of urea (9.5 M) provides unusual stability to the protein with no more structural loss and transition to a completely unfolded state.
KEY WORDS: stem bromelain, urea denaturation, protein stability