pH-Dependent Urea-Induced Unfolding of Stem Bromelain: Unusual Stability against Urea at Neutral pH

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B. Ahmad, G. M. Rathar, A. Varshney, and R. H. Khan^*

Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh 202002, India; fax: +91(571)272-1776; E-mail: rizwanhkhan@hotmail.com; rizwanhkhan1@yahoo.com

^* To whom correspondence should be addressed.

Received March 25, 2009; Revision received April 20, 2009

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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 C_m (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.

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KEY WORDS: stem bromelain, urea denaturation, protein stability

DOI: 10.1134/S0006297909120062

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