COOPERATIVITY IN THE UNFOLDING TRANSITIONS OF CYSTEINE PROTEINASES - CALORIMETRIC STUDY OF THE HEAT DENATURATION OF CHYMOPAPAIN AND PAPAIN

Differential scanning calorimetry (DSC) was employed to study the ther mal unfolding of chymopapain (EC 3.4.22.6) and papain (EC 3.4.22.2), t wo highly homologous cysteine proteinases from Carica papaya. Under al l pH conditions used, both enzymes showed irreversible thermal denatur ation. However, results from experiments performed at two different sc anning rates suggest that interpretation of data in terms of equilibri um thermodynamics is not unreasonable. For papain, the ratio of calori metric (DELTAH(cal)) to van't Hoff (DELTAH(vH)) enthalpies approximate d to 2.0. This value indicates that papain domains unfold almost indep endently, as it has been reported previously. In contrast, chymopapain displayed a more cooperative behavior with a DELTAH(cal) to DELTAH(vH ) ratio of 1.3-1.4. DSC curves were analyzed in terms of a mechanism t hat includes domain-domain interactions. The results showed a negligib le interdomain free energy in the case of papain, but a significant va lue of approx. 1.0 kcal/mol (1 cal = 4.184 J) for chymopapain. These t wo proteins also differed in the unfolding heat-capacity change, DELTA C(p), which suggests that their native structures bury different amoun ts of nonpolar surface area.