Noncovalent enzyme immobilization methods

In summary, enzyme immobilization is extremely important in the scale-up of many biocatalytic processes. The preferred method for pharmaceutical production involves covalent binding through cross-linking or attachment to a support. Noncovalent attachment is less attractive, but it is heavily utihzed owing to the commercial availabihty of industrial quantities of some enzymes immobilized using this technique. 

Most biocatalytic conversions are performed with the enzyme immobilized in the microreactor. Miyazaki et al. [426] developed a simple noncovalent immobilization method for His-tagged enzymes on a microchannel surface. These enzymes contain a polyhistidine-tag motif that consists of at least six histidine residues, often located at the N- or C-terminus. The H is-tag has a strong affinity for nickel and can be reversibly immobilized by a nickel-nitrilotriacetic acid (Ni-NTA) complex (Scheme 4.103), a strategy commonly used in affinity chromatography. 

The adsorption of an enzyme onto a support or film material is the simplest method of obtaining an immobilized enzyme. Basically, the enzyme is attached to the support material by noncovalent linkages and does not require any preactivation step of the support. The interactions formed between the enzyme and the support material will be dependent on the existing surface chemistry of the support and on the type of amino acids exposed at the surface of the enzyme molecule. Enzyme immobilization by adsorption involves, normally, weak interactions between the support and the enzyme such as ionic or hydrophobic interactions, hydrogen bonding, and van der Waals forces (see Figure 44.1). ... 

Conditions for achieving efficient DET via enzyme immobilization are dictated partly by materials architecture. Enzyme immobilization techniques may include nonspecific adsorption, covalent linkage, entrapment in conductive polymeric films, association with metal colloids, and encapsulation within porous matrices (see Chapter 11). The simplest method is nonspecific adsorption, but control is limited various noncovalent interactions will yield different orientations of the redox center with respect to the electrode interface and, as a result, inefficient DET. 

There are distinct methods to improve the enzyme stability either by simple coating of enzymes by ILs or by noncovalent adsorption or by covalent linking of the enzymes with the solid supports such as polymers, nanoparticles, carbon nanotubes, and encapsulation in hydrogels. One of the interesting and new features of ILs as an immobilization support for enzymes evolved out of its incompetence for retaining... 

The final common method for employing enzymes in biocatalysis involves their prior immobilization on a solid support by covalent or noncovalent bonds. In many cases, this modification deaeases their catalytic effidendes, and it can also introduce mass transport limitations. Minimizing both of these issues by optimizing both the nature of the solid support and the enzyme/support connection is an area of intensive current research. Some enzymes are available commerdally in immobilized form. The best-known example is Novoz5mi 435, which is a trade name for Candida antarctica lipase B adsorbed on a polymeric support. 

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