Immobilization of enzymes
has been achieved by intermolecular cross-linking of the protein, either
to other protein molecules or to functional groups on an insoluble support
matrix.. Cross-linking an enzyme to itself is both expensive and insufficient,
as some of the protein material will inevitably be acting mainly as a support.
This will result in relatively low enzymatic activity. Generally, cross-linking
is best used in conjunction with one of the other methods. It is used mostly
as a means of stabilizing adsorbed enzymes and also for preventing leakage
from polyacrylamide gels.
Since the enzyme is covalently
linked to the support matrix, very little desorption is likely using this
method. Marshall (1973), for example, reported that carbamy phosphokinase
cross-linked to alkyl amine glass with glutaraldehyde lost only 16% of
its activity after continuous use in a column at room temperature for fourteen
The most common reagent
used for cross-linking is glutaraldehyde. Cross-linking reactions are carried
out under relatively severe conditions. These harsh conditions can change
the conformation of active center of the enzyme; and so may lead to significant
loss of activity.
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