Role of Hydrodynamic Shear on Activity and Structure of Proteins
55
deactivation of cellulase by shear becomes evident when digestion of the more
crystalline forms of cellulose is performed and this might be one of the reasons
for the decrease in reaction rate during the enzymatic hydrolysis of cellulose.
In a further study on cellulase enzyme, Kim et al. [31] recirculated the
enzyme through a capillary tubing in the presence of different stabilizers. They
also investigated the effect of air-liquid interracial forces by passing the enzyme
solution through tubing from which all air bubbles had been removed. Under
these conditions cellulase was not inactivated even after 4 h at a shear rate of
850 s- 1. However, in the presence of air in the tubing, a situation similar to those
under normal operating conditions was observed and there was a 60% loss in
activity. The deactivation of the enzyme also showed dependence on the protein
concentration present. Various stabilization agents were used in the presence of
air-liquid interface and it was found that Zonyl, Triton X-100 and bovine serum
albumin were able to decrease the deactivation significantly whereas sodium
dodecyl sulphate and lysozyme were not effective. The effect of these additives is
mainly due to their ability to minimize the concentration of enzyme present at
the air-liquid interface. Studies carried out with an extremely shear sensitive
enzyme viz. ribulose-l,5-biphosphate carboxylase/oxygenase showed that bovine serum albumin was able to minimise shear damage in the presence of an
air-liquid interface. This enzyme undergoes deactivation even under very mild
conditions such as pipetting or vortexing in the laboratory [32]. The enzyme
also shows spontaneous deactivation on standing. These effects were examined
by subjecting the enzyme to shear under conditions similar to those used by
Charm and Wong [27]. The enzyme showed deactivation at a relatively weak
shear stress corresponding to an average shear rate x time of 105 and this
deactivation was significantly decreased by the addition of bovine serum albumin (1 mgml- l) [33].
From this discussion it becomes clear that any unit operation which requires
a protein solution to be passed through a tubing has the potential to cause
deactivation of the protein. The denaturation of proteins under these conditions
is pronounced in the presence of an air-liquid interface. The air-liquid interface
may exist due to the presence of small amounts of air in the form of bubbles
being entrapped within the equipment or the solution itself. Addition of certain
surface active agents such as Zonyl, Triton X-100, and serum albumin can
alleviate the problem of shear induced denaturation of proteins especially in the
presence of entrapped air. However, systematic work is needed for understanding the mechanism of action of stabilization of these additives. Furthermore,
a systematic procedure is also required for the selection of the additive and its
concentration for a given application.
3.2 Concentric Cylinder Viscometer
Another device used for generating defined shear stresses is a viscometer consisting of two cylinders placed concentrically. The flow in this device is of couette
55
deactivation of cellulase by shear becomes evident when digestion of the more
crystalline forms of cellulose is performed and this might be one of the reasons
for the decrease in reaction rate during the enzymatic hydrolysis of cellulose.
In a further study on cellulase enzyme, Kim et al. [31] recirculated the
enzyme through a capillary tubing in the presence of different stabilizers. They
also investigated the effect of air-liquid interracial forces by passing the enzyme
solution through tubing from which all air bubbles had been removed. Under
these conditions cellulase was not inactivated even after 4 h at a shear rate of
850 s- 1. However, in the presence of air in the tubing, a situation similar to those
under normal operating conditions was observed and there was a 60% loss in
activity. The deactivation of the enzyme also showed dependence on the protein
concentration present. Various stabilization agents were used in the presence of
air-liquid interface and it was found that Zonyl, Triton X-100 and bovine serum
albumin were able to decrease the deactivation significantly whereas sodium
dodecyl sulphate and lysozyme were not effective. The effect of these additives is
mainly due to their ability to minimize the concentration of enzyme present at
the air-liquid interface. Studies carried out with an extremely shear sensitive
enzyme viz. ribulose-l,5-biphosphate carboxylase/oxygenase showed that bovine serum albumin was able to minimise shear damage in the presence of an
air-liquid interface. This enzyme undergoes deactivation even under very mild
conditions such as pipetting or vortexing in the laboratory [32]. The enzyme
also shows spontaneous deactivation on standing. These effects were examined
by subjecting the enzyme to shear under conditions similar to those used by
Charm and Wong [27]. The enzyme showed deactivation at a relatively weak
shear stress corresponding to an average shear rate x time of 105 and this
deactivation was significantly decreased by the addition of bovine serum albumin (1 mgml- l) [33].
From this discussion it becomes clear that any unit operation which requires
a protein solution to be passed through a tubing has the potential to cause
deactivation of the protein. The denaturation of proteins under these conditions
is pronounced in the presence of an air-liquid interface. The air-liquid interface
may exist due to the presence of small amounts of air in the form of bubbles
being entrapped within the equipment or the solution itself. Addition of certain
surface active agents such as Zonyl, Triton X-100, and serum albumin can
alleviate the problem of shear induced denaturation of proteins especially in the
presence of entrapped air. However, systematic work is needed for understanding the mechanism of action of stabilization of these additives. Furthermore,
a systematic procedure is also required for the selection of the additive and its
concentration for a given application.
3.2 Concentric Cylinder Viscometer
Another device used for generating defined shear stresses is a viscometer consisting of two cylinders placed concentrically. The flow in this device is of couette