conformational stability is less, and if the adsorbent is more hydrophobic,
presumably because the driving force for conformational change is greater.
It is sometimes observed that adsorption from a more dilute enzyme
solution leads to more inactivation. The explanation may be that at low
concentration adsorption is slow, allowing adsorbed molecules to expand
laterally, which implies conformational change. If adsorption is fast, a
densely packed adsorbed layer is rapidly formed, which would prevent
lateral expansion. In agreement with this, it has been observed that some
proteins do not greatly change conformation when merely adsorbing onto
an air–water interface, but when the air–water surface is expanded, for
instance by deforming an air bubble, considerable change occurs. Beating
air into a protein solution can therefore cause denaturation.
It has further been observed for several enzymes that adsorption onto
an oil–water interface causes complete inactivation, whereas only partial
inactivation may occur due to adsorption onto an air–water surface. The
reason may be that hydrophobic segments of the molecule can penetrate
into an oil phase, but not into air. This would be because the net attractive
energy between these segments and oil can be greater than that between
segments and water, whereas the attractive energy between any group of a
protein and air will be virtually zero. This must cause a greater driving force
for loss of native configuration at the oil–water interface. A fairly stable
enzyme like lysozyme, which can regain activity after various unfolding
treatments at low temperature, does not regain it after adsorption onto oil
droplets, even at its isoelectric pH. This leads to the important conclusion
that more than one unfolded state can exist, and that some of these states
permit return to the native state, whereas others do not.
- Shear stress. It has been observed that some enzymes under some
conditions (such as the presence of specific solutes) show inactivation when
the solution is subjected to simple shear flow for a considerable time,
especially when the temperature is not much below the denaturation
temperature. The extent of inactivation then is proportional to the product
of shear rate and treatment time. Most workers agree that the shearing
stresses applied (for instance 1 Pa at a shear rate of 1000 s^1 and a viscosity
of 1 mPa?s) are far too small to affect protein conformation. In some cases,
denaturation at the air–water interface may have occurred, but in other
cases this possibility has been ruled out. There is no generally accepted
explanation of the effect.