Chemical methods

• Chemical methods of enzyme immobilization

involve binding enzymes to a solid support
through chemical reactions.
• These methods provide a strong and stable
attachment of enzymes to the support material,
allowing for improved enzyme stability and
reusability.
Covalent Bonding
• Enzymes can be covalently bonded to a solid support using
chemical reactions.
• Functional groups on the enzyme (e.g., amino groups on lysine
residues) can react with functional groups on the support material
(e.g., activated carboxyl or aldehyde groups).
• The covalent binding basically takes place by activation of the
support via adding reactive molecules then altering the polymer’s
backbone to activate the whole matrix.
• The hydrophilic polysaccharide polymers are the most commonly
used supports in this strategy.
 Cross linking
• Cross-linking immobilization is a strategy where
enzymes are interconnected through covalent
bonding without carriers.
• The intermolecular cross-linking is accomplished
through the presence of linker agents, which are
used as bridges between two adjacent enzyme
molecules.
• Cross-linking immobilization delivers a robust
connection between the enzymes, leading to
superior stabilities.
• Aggregates of cross-linked enzymes are formed
through straightforward enzyme precipitation in
an aqueous media, which produces physical
aggregates composed of the protein particles.
Affinity Binding
• This method utilizes the specific affinity between a
ligand (e.g., antibodies, lectins) and a corresponding
receptor on the enzyme.
• The ligand is immobilized on a solid support, and the
enzyme binds selectively to it.
• Affinity binding offers highly specific immobilization,
but it may require specialized ligands.
Ionic Binding
• This method relies on electrostatic interactions between the
enzyme and a support material.
• In this technique, the enzyme and the support material have
opposite electrical charges, leading to the binding of the
enzyme to the support material through electrostatic forces.
• The support material used in ionic binding typically has
charged functional groups.
• The enzyme is usually in a buffered solution, and its charge
depends on the pH of the solution.
• When the pH of the enzyme solution matches the pH of the
support material, the enzyme and the support material will
have opposite charges.
• fter adsorption, the immobilized enzyme is typically washed
to remove any unbound or weakly bound enzyme molecules.