ABSTRACT
We studied the irreversible thermal denaturation of chymopapain, a papain-related cysteine proteinase. It was found that this process follows simple first-order kinetics under all conditions tested. Rate constants determined by monitoring ellipticity changes at 220 or 279 nm are essentially identical, indicating that denaturation involves global unfolding of the protein. Enthalpies (DeltaH(double dagger)) and entropies (DeltaS(double dagger)) of activation for unfolding were determined at various pH values from the temperature dependence of the rate constant. In the pH range 1.1-3.0, a large variation of both DeltaH(double dagger) and DeltaS(double dagger) was observed. For the few proteins studied so far (lysozyme, trypsin, barnase) it is known that activation parameters for unfolding vary little with pH. It is proposed that this contrasting behavior of chymopapain originates from the numerous ion pairs - especially those with low solvent accessibilities - present in its molecular structure. In contrast, fewer, more exposed ion pairs are present in the other proteins mentioned above. Our results were analyzed in terms of differences in the protonation behavior of carboxylic groups between the transition (TS) and native (N) states of the protein. For this purpose, a model of independently titrating sites was assumed, which explained reasonably well the pH dependence of activation parameters, as well as the protonation properties of native chymopapain. According to these calculations, pK values of carboxyls in TS are shifted 0.6-0.9 units upwards with respect to those in N. In addition, some groups in TS appear to be protonated with unusually large enthalpy changes.
Subject(s)
Chymopapain/chemistry , Enzyme Activation , Enzyme Stability , Protein Folding , Circular Dichroism , Cysteine Endopeptidases/chemistry , Hydrogen-Ion Concentration , Kinetics , Plant Proteins/chemistry , Protein Denaturation , Protons , ThermodynamicsABSTRACT
An experience of feeding healthy premature infants (< 1,800 g) on their mother's milk is communicated. Feeding starts at an early time (15 hours); achieving maximum volumes at the 12th day. Loss of weight is 10% less than the birth weight and recovery occurs at the 12th day. In this way, a postnatal speed growth of 26 g/day can be obtained; which is similar than that observed during the third of the pregnancy. 75% of the calories provided to more of the 66% of the infants attended by our Unity, is based on their own mother's milk at the time of discharge.
