Immobilized Enzymes (Quiz 1) Flashcards
The restriction of enzyme mobility in a fixed space
Enzyme Immobilization
Immobilization of enzymes provides Important advantages, (4)
- Enzyme Reutilization
- Elimination of Enzyme Recovery
- Purification processes
- May provide a better environment for enzyme activity.
Since some of the intracellular enzymes are ________ Immobilized enzymes provide a model system to __________ the action of some membrane-bound intracellular enzymes.
membrane bound
mimic and understand
Product purity is usually improved, and effluent handling problems are minimized by ________
immobilization
Methods of Immobilization (2)
- Entrapment
- Surface Immobilization
is the physical enclosure of enzymes in a small space
Entrapment
Two Major Methods of Entrapment
- Matrix entrapment
- Membrane entrapment, including microencapsulation
________ used for enzyme immobilization are usually polymeric materíals such as Ca-alginate,
agar, k- carrageenin, polyacrylamide, and
collagen.
- However, some solid matrices such as activated
carbon, porous ceramic, and diatomaceous earth
can also be used for this purpose.
Matrices
can be a particle, a membrane, or a fiber.
Matrix
______________ of enzymes is possible;
Ex.
- hollow fiber units have been used to entrap an enzyme solution between thin, semipermeable membranes.
- Membranes of nylon,
cellulose, polysulfone, and polyacrylate are commonly used. - Configurations, other than hollow fibers, are possible, but in all cases a _______ is used to retain high-molecular weight compounds (enzyme), while allowing small molecular- weight compounds (substrate or products) access to the enzyme
Membrane entrapment
semipermeable membrane
A special form of membrane entrapment.
- In this technique, microscopic
hollow spheres are formed. - The spheres contain the enzyme solution, while the sphere is enclosed within a porous membrane.
- The membrane can be polymeric or an enriched interfacial phase formed around a microdrop.
Microencapsulation
enzyme entrapment may have its inherent problems, such as: (4)
- Enzyme leakage into solution
- Significant diffusional limitations
- Reduced Enzyme Activity and Stability
- lack of control of Microenvironmental conditions
____________ can be overcome by reducing the MW cutoff of membranes or the pore size of solid matrices.
Enzyme leakage
_________ can be eliminated by reducing the particle size of matrices and/or capsules.
Diffusion limitations
_________________ are due to unfavorable microenvironmental conditions, which are difficult to control.
However, by using different matrices and chemical ingredients, by changing processing conditions, and by reducing particle or capsule size, more favorable microenvironmental conditions can be obtained.
Reduced enzyme activity and stability
____________ is usually less significant in microcapsules as compared to gel beads.
Diffusion barrier
The two major types of immobilization of enzymes on the surfaces of support materials are:
- adsorption
- covalent binding
The active site of the adsorbed enzyme is usually unaffected, and nearly full activity is retained upon ________.
adsorption
Type of Surface Immobilization:
is the attachment of enzymes on the surfaces of support particles by weak physical forces, such as van der Waals or dispersion forces.
Adsorption
_________ of enzymes is a common problem, especially in the presence of strong hydrodynamic forces, since binding forces are weak.
desorption
Adsorption of enzymes may be stabilized by cross-linking with ____________. Its treatment can denature some proteins.
glutaraldehyde
Support materials used for enzyme adsorption can be:
- _____________, such as alumina, silica, porous glass, ceramics, diatomaceous earth, clay
- _________, or ________, such as
cellulose (CMC, DEAE-cellulose), starch, activated carbon - ___________, such as
Amberlite, Sephadex, and Dowex.
- inorganic materials
- bentonite or organic materials
- ion-exchange resins
________________ is the retention of enzymes on support surfaces by ______ bond formation.
Covalent Binding
Covalent
Enzyme molecules bind to support material via certain functional groups,
such as ________, _______, ________, and ________ groups.
These functional groups must not be in the active site
- amino
- carboxyl
- hydroxyl
- sulfhydryl
One common trick Is to block the active site by ______ the enzyme solution with a competitive inhibitor prior to covalent binding.
flooding
Functional groups on support material are usually activated by using chemical reagents, such as:
- cyanogen bromide
- carbodiimide
- glutaraldehyde
Two major criteria used in the selection of support material are:
- the _____ of the support material, which is a function of charge density, functional groups, porosity, and hydrophobicity of the support surface.
- _____ and ________ of enzymatic activity, which is a function of functional groups on support material and microenvironmental conditions.
- binding capacity
- stability and retention
If immobilization causes some _______ changes on the enzyme, or if reactive groups on the active site of the enzyme are involved in binding, a ______ in enzyme activity can take place upon immobilization.
conformational
loss
Usually, immobilization results in a loss in enzyme activity and stability.
However, in some cases,
Immobilization may cause an increase in enzyme activity and stability due to more favorable
_________. Because enzymes often have more than one _______ that can bind the surface, an immobilized enzyme preparation may be very __________.
microenvironmental conditions
functional site
heterogeneous
Even when binding does not _____ enzyme structure, some enzyme can be bound with the active site oriented away from the _____ and toward the ______, decreasing the access of the substrate to the enzyme. __________ of activity varies with the method used.
alter
substrate solution
support surface
Retention
