Multiple intermediate conformations of jack bean urease at low pH: anion-induced refolding.

Structural and functional characteristics of jack bean urease (JBU), a hexameric enzyme having identical subunits, were investigated under neutral as well as acidic conditions by using CD, fluorescence, ANS binding and enzyme activity measurements. At low pH and low ionic strength, JBU exists in a partially unfolded state (U(A)-state), having predominantly beta structure and no tertiary interactions along with a strong ANS binding. Addition of salts like NaCl, KCl and Na(2)SO(4) to the U(A)-state induces refolding resulting in structural propensities similar to that of native hexamer. Moreover, at low concentrations, GuHCl behaves like an anion by inducing refolding of the U(A)-state. The anion-induced refolded state (I(A)-state) is more stable than U(A)-state and the stability is nearly equal to that of the native protein against chemical-induced and thermal denaturation. Overall, these observations support a model of protein folding for a multimeric protein where certain conformations (ensembles of substates) of low energy prevail and populated under non-native conditions with different stability.