There are two forms of rabbit muscle adenylate kinase (AK) with different 8-anilino-1-naphthalenesulfonic acid (ANS) binding properties in equilibrium solution. One form (about 70%, denoted N1) binds rapidly with ANS, whereas the other (about 30%, denoted N2) does not. Furthermore, native forms of AK should adopt different conformations for binding with substrates and products, which should be pre-existing for performing its catalytic function. The present experiments demonstrate both forms of AK distinguished by ANS probe are active. The activity of N2 is about 0.8 fold higher than N1 and shows higher susceptibility to proteolysis by trypsin. This means that the native state of AK might be an ensemble of kinetically attainable conformers and the energy landscapes of AK folding should be rugged with more than one local minimum.
