Physical Chemistry of Foods

which does not have a double-bond character, implying that rotation about
the 22 CO 22 N<bond is possible; trans and cis conformations have about
equal stability in this case. The change involves an activation enthalpy of
about 85 kJ?mol
^1. The relaxation time thus is about 20 min at 0 8 C and 1 s
at 70 8 CðQ 10 & 3 Þ. This implies that these peptide bonds will have about
equal amounts of trans and cis at high temperature in an unfolded chain;
rapid cooling may then readily cause locking of cis bonds. In such a case,
refolding would not be in the native conformation, but slow renaturation
may occur after cooling. After all, some protein molecules, though it may be
a very small number, will be in the unfolded state, even at the optimum
temperature for stability.

3. Reshuffling (scrambling) of sulfur bridges. This occurs if the
   molecule is unfolded and contains a free 22 SH, of which at least a little is
   present in its ionized form, i.e. at pH>about 6. The reaction occurring is
   schematically

Ra 22 S 22 S 22 RbþRc 22 S
/


?

Ra 22 S 22 S 22 RcþRb 22 S

Interchange of disulfides can also occur among different protein molecules,
producing molecular aggregates. These reactions occur faster at higher
temperature and higher pH. At very high pH, 22 S 22 S 22 bridges are in fact
weak bonds that readily break.

4. Deamidation. At high temperatures, the residue Asn may show the
   reaction

22 CONH 2 þH 2 O 
? 22 COOHþNH 3

thereby forming Asp. It may also happen with Gln, giving Glu, though to a
far smaller extent. These reactions occur faster at a lower pH. The changes
would generally lead to a nonnative conformation upon refolding.

5. At very high temperatures variouscross-linkingreactions between
   side groups are possible. Moreover, peptide bonds may even be hydrolysed,
   depending on pH.


6. The absence ofchaperonins, as discussed above.

Denaturing Agents. As mentioned, several agents or conditions
can cause denaturation. They may be categorized as follows:

1. High or low temperature. Conformational stability as a function of
   temperature is discussed in the previous section; see especially Figure 7.5. By
   and large, unfolding at low temperature occurs because hydrophobic bonds
   then are very weak or even repulsive. At high temperature, the increased
   effect of the conformational entropy becomes overriding. At high
   temperature, irreversible changes in protein configuration may well occur,