Sequential unfolding of papain in molten globule state.

Papain exhibits the characteristics of molten globule under acidic conditions as seen by circular dichroism, fluorescence and ANS binding. Between pH 2.0-2.5 the protein exhibits substantial secondary structure as indicated by far-UV CD spectrum but loses the persistent tertiary interactions of the native state. Enhanced binding of ANS to the state at pH 2.0 in relation to the native and unfolded states at neutral pH indicates a considerable exposure of aromatic side chains. Temperature and guanidine hydrochloride induced unfolding of papain in this state is noncooperative and the transition curves are biphasic in nature. As papain molecule consists of two domains, the results suggest that the domains unfold independently and sequentially.