BIOTECHNOLOGY - V. H. Herasymenko - 2006

Part II. Special Biotechnologies

Chapter 7. BIOTECHNOLOGY OF THE PRODUCTION AND APPLICATION OF IMMOBILIZED PREPARATIONS

7.3. ENZYME IMMOBILIZATION. OBJECTIVES OF IMMOBILIZATION

Enzyme Immobilization refers to the incorporation of an enzyme into a distinct isolated phase separated from the free solution phase, yet capable of exchanging substrate or effector molecules present in the free solution phase. In other words, immobilization is the inclusion of an enzyme into an environment where only a limited portion of the total volume is accessible to it.

An immobilized enzyme (from Lat. immobilis — motionless) is an enzyme attached by one means or another to an inert, insoluble support or carrier (matrix) while partially or completely retaining its catalytic activity.

An enzyme placed within a mesh-like matrix through which low-molecular-weight substrates and reaction products can freely penetrate, while enzyme protein macromolecules are excluded, is also considered immobilized.

Immobilization restricts molecular mobility and anchors the Structure in such a way that the Active Site of the enzyme preserves its catalytic activity over an extended period of time.

The objective of immobilization is to enable The Use of enzyme preparations as industrial catalysts in biotechnological processes for producing biotech products, as well as for analytical and therapeutic purposes.

The Combination of an enzyme with a carrier led to the creation of heterogeneous biocatalysts, for which the term "immobilized Enzymes" was officially adopted at the First Engineering Enzymology Conference in Henniker (USA) in 1971. In literature, one may still encounter other terms, such as "insoluble enzymes," "matrix-bound enzymes," etc., which share the exact same meaning: these are enzyme preparations bound to insoluble Supports.

However, METABOLISM/2.html">THE CONCEPT OF immobilization should be viewed more broadly, encompassing any restriction on the spatial freedom of movement of protein molecules (or their fragments). In addition to binding to an insoluble support, this can be achieved through intra- or intermolecular cross-linking of protein molecules using low-molecular-weight bifunctional Reagents, or by attaching the enzyme to a soluble polymer. Such preparations are referred to as enzymes modified with "cross-linking" or polymeric reagents, respectively.

Immobilized enzyme preparations offer A number of significant advantages for applied use compared to their native precursors.

First, upon immobilization, enzymes transition from homogeneous catalysts (which exist in the same phase as the reaction substrates) to heterogeneous catalysts (which form a distinct phase separated from the reactants). Consequently, the enzyme and the reactants can be separated. This makes it possible to: a) stop the reaction at the desired moment; b) regenerate the enzyme upon reaction completion and reuse the catalyst; c) obtain a product uncontaminated by the enzyme, which is crucial in food and pharmaceutical manufacturing.

Second, the use of heterogeneous catalysts allows the enzymatic process to be carried out continuously, for example, in packed-bed columns, and makes it possible to regulate both the catalyzed reaction rate and product yield by adjusting the flow rate.

Third, immobilization or modification of an enzyme facilitates the targeted alteration of the catalyst's properties, including its Specificity, the dependence of its catalytic activity on pH, Ionic strength, Temperature, and other environmental parameters, while also increasing its resistance (or stability) to various denaturing factors by 100 to 1000 times.

Fourth, enzyme immobilization provides The ability to regulate their catalytic activity by altering The properties of the carrier.

The advent of this new Class of bio-organic catalysts—immobilized enzymes—has made it possible to establish large-scale industrial manufacturing of biotechnological products that is amenable to automation.

The immobilization method also makes it possible to obtain preparations with prolonged and targeted action for therapeutic Applications in human and veterinary medicine, as well as in animal husbandry to improve Digestion, enhance the utilization of feed nutrients in animals, and increase their productivity.