Abstract
Differential scanning calorimetry (DSC) was employed to study the thermal unfolding of chymopapain (EC 3.4.22.6) and papain (EC 3.4.22.2), two highly homologous cysteine proteinase from Carica papaya. Under all pH conditions used, both enzymes showed irreversible thermal denaturation. However, results from experiments performed at two different scanning rates suggest that interpretation of data in terms of equilibrium thermodynamics is not unreasonable. For papain, the ratio of calorimetric ( ΔH cal) to van't Hoff ( ΔH vH) enthalpies approximated to 2.0. This value indicates that papain domains unfold almost independently, as it has been reported previously. In contrast, chymopapain displayed a more cooperative behavior with a ΔH cal to ΔH vH ratio of 1.3–1.4. DSC curves were analyzed in terms of a mechanism that includes domain-domain interactions. The results showed a negligible interdomain free energy in the case of papain, but a significantly value of approx. 1.0 kcal/mol (1 cal = 4.184 J) for chymopapain. These two proteins also differed in the unfolding heat-capacity change, ΔC p, which their native structures bury different amounts of nonpolar surface area.
Published Version
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