Enzyme binding capacity

Enzyme Binding Capacity Determination. Two steps or stages are involved in formation of a stable collagen/enzyme complex. 

Unless otherwise stated, the enzyme binding capacity of membraneous collagen was measured by the equilibrium sorption method. 

Chemical Modification of Collagen- The Effect of Carbamyla-tion on the Enzyme-Binding Capacity of Membraneous Collagen. 

As previously discussed, the decrease in enzyme binding capacity of the chemically modified membrane implies that the complexation mechanism involves ionic interactions of lysyl E-amino groups of collagen with enzymic amino acid side chains as a principle step in the formation of a stable network of phyico-chemical bonds. However, part of the effect of carbamylation could be due to a reduction in the average binding constant (K0) of all the sites rather than the complete blocking of active sites. 

An additional problem arises when the exchange processes are rate-limited. This may be caused by enzymes that become saturated when all their active sites are occupied by the drug, or it may be due to adsorbing proteins that have a limited binding capacity. In such cases, one obtains a type of Michaelis-Menten kinetics of the form ... 

In order to compare the specific activity of plant-derived C5-1 to that of the hybridoma-derived antibody, the antigen-binding capacity of antibodies produced in each system was assayed by enzyme-linked immunosorbent assay (ELISA). As shown in Table 1.2, antibodies from both sources demonstrated similar binding characteristics against human IgGs [8]. Furthermore, the stability of alfalfa-derived C5-1 in the blood stream of Balb/c mice was comparable to that of the hybridoma-derived IgG [8]. 

Adults require 1-2 mg of copper per day, and eliminate excess copper in bile and feces. Most plasma copper is present in ceruloplasmin. In Wilson s disease, the diminished availability of ceruloplasmin interferes with the function of enzymes that rely on ceruloplasmin as a copper donor (e.g. cytochrome oxidase, tyrosinase and superoxide dismutase). In addition, loss of copper-binding capacity in the serum leads to copper deposition in liver, brain and other organs, resulting in tissue damage. The mechanisms of toxicity are not fully understood, but may involve the formation of hydroxyl radicals via the Fenton reaction, which, in turn initiates a cascade of cellular cytotoxic events, including mitochondrial dysfunction, lipid peroxidation, disruption of calcium ion homeostasis, and cell death. 

Cows and calves fed low-zinc diets of 25 mg Zn/kg ration showed a decrease in plasma zinc from 1.02 mg/L at start to 0.66 mg/L at day 90 cows fed 65 mg Zn/kg diet had a significantly elevated (1.5 mg Zn/L) plasma zinc level and increased blood urea and plasma proteins (Ram-achandra and Prasad 1989). Biomarkers used to identify zinc deficiency in bovines include zinc concentrations in plasma, unsaturated zinc-binding capacity, ratio of copper to zinc in plasma, and zinc concentrations in other blood factors indirect biomarkers include enzyme activities, red cell uptake, and metallothionein content in plasma and liver (Binnerts 1989). 

As already discussed, a covalent immobilization can be performed via different chemical moieties on the protein surface. Because of that, protein molecules are immobilized in random orientation with at least one, but often several, covalent bonds to the matrix. As a result, the active site might be oriented toward the matrix surface and its accessibility to the substrate molecule hence significantly reduced. This results in a decrease of biological activity and consequently in lower binding capacity or decrease of reaction rate in the case of enzymes. 

The hydrolytic activities of the intact enzymes were comparable, but CBH I was much more sensitive to product (cellobiose) inhibition. Both core enzymes exhibited a strongly reduced activity (50-90%) which was correlated with the absence of the binding domain and their consequent lower binding capacity on Avicel. The activities of CBH I and Core I on amorphous cellulose were, however, comparable. 

The ability of proteins to exist in different conformations is termed allostery (see 2.2). Allosteric enzymes can assume various conformations which differ in catalytic activity and/or substrate binding capacity. 

Water Uptake and Retention. The water binding capacity of soy protein isolate can be increased by treatment with neutral fungal protease (20). Since the number of free amino and carboxyl groups increases as a result of digestion and because moisture uptake by proteins is proportional to the number of ionic groups present ( ), it is not surprising that moisture uptake is increased by enzyme treatment. 

Surugiu et al. [67] have introduced an Enzyme Immuno-Like Assays (EzILA) for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The label was a 2,4-D conjugate with the tobacco peroxidase (TOP) enzyme, which allows for both colorimetric and chemiluminescent detection. In this case, the polymer imprinted with 2,4-D was synthesized in the form of microspheres. In contrast, despite its higher binding capacity for radiolabeled 2,4-D, a conventional MIP prepared by bulk polymerization showed only weak binding of the 2,4-D-TOP tracer. 

Natural supports (agarose, dextran, cellulose, porous glass, silica, the optical fiber itself or alumina) and synthetic resins (acrylamide-based polymers, methacrylic acid-based polymers, maleic anhydride-based polymers, styrene-based polymers or nylon, to name a few) have been applied for covalent attachment of enzymes. These materials must display a high biocatalyst binding capacity (as the linearity and the limit of detection of the sensing layers will be influenced by this value), good mechanical and chemical stability, low cost, and ease of preparation. 

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