Immobilization of Enzymes by Selective Adsorption on Biotinylaminopropyl Celite or Glass

Use of immobilized enzymes in bioprocesses offers many advantages, including greater productivity because the same enzyme molecules can be used over a long period of time, there is more precise control of the extent of reaction, there is the capability of automation and continuous operation, and the elimination of the requirement of a downstream enzyme inactivation step (1 ). The ease and cost of immobilized enzyme preparation has limited commercial application of enzyme bioreactor processes. In this chapter a method for immobilization of enzymes using the high affinity, bioselective interaction between biotin and avidin is described. Avidin, which is a tetrameric protein, binds four biotin molecules with a dissociation constant of about 10⁻¹⁵ (2 ) thus, it represents one of the strongest noncovalent interactions known. Vis-a-vis adsorption by ion-exchange interaction, the avidin-biotin interaction is very specific and much stronger; therefore, enzyme leaking from the support is minimal, resulting in greater operational stability (3 –9 ). Furthermore, the biotin molecule is very robust; consequently, bioreactors can be repeatedly regenerated simply by desorption of inactive enzyme in a chaotropic solvent, followed by selective adsorption of avidin and finally, fresh biotinylated enzyme.