Horseradish peroxidase isolation

Other specific discovery assays have been used such as differential inhibition of a vancomycin resistant S. aureus strain and its susceptible parent, and an assay based on antagonism of the antibacterial activity by N,A/-diacetyl-L-Lys-D-Ala-D-Ala [24570-39-6] a tripeptide analogue of the dalbaheptides receptor. AppHcation of this latter test to 1936 cultures (90) led to the isolation of 42 dalbaheptides, six of which, including kibdelin (Table 3), parvodicin (Table 3), and actinoidin A2 (68) were novel. A colorimetric assay based on competition between horseradish peroxidase bound teicoplanin and the... 

Attachment of Hydroxycinnamic Acids to Structural Cell Wall Polymers. Peroxidase mediation may also result in binding the hydroxycinnamic acids to the plant cell wall polymers (66,67). For example, it was reported that peroxidases isolated from the cell walls of Pinus elliottii catalyze the formation of alkali-stable linkages between [2-14C] ferulic acid 1 and pine cell walls (66). Presumably this is a consequence of free-radical coupling of the phenoxy radical species (from ferulic acid 1) with other free-radical moieties on the lignin polymer. There is some additional indirect support for this hypothesis, since we have established that E-ferulic acid 1 is a good substrate for horseradish peroxidase with an apparent Km (77 /tM), which is approximately one fifth of that for E-coniferyl alcohol (400 /iM) (unpublished data). 

Wuhrer, M., Balog, C. I., Koeleman, C. A., Deelder, A. M., and Hokke, C. H., New features of site-specific horseradish peroxidase (HRP) glycosylation uncovered by nano-LC-MS with repeated ion-isolation/ fragmentation cycles, Biochimica et Biophysica Acta 1723(1-3), 229-239, 2005. 

Pitkanen et al. [51] reported the isolated bovine RPE-choroid was up to 20 times more permeable to lipophilic than hydrophilic beta-blockers. Furthermore, the in vitro permeability of bovine RPE-choroid to hydrophilic compounds and macromolecules was 10 to 100 times less compared to sclera, whereas the permeability for lipophilic molecules was in the same range for both tissues. The isolated bovine RPE-choroid also exhibited differential permeation by molecular weight and Stokes radius. The permeation rate of 4, 10, and 20 kDa FITC dextrans was moderate compared to a good permeation rate for the 376 Da carboxyfluor-escein and a poor penetration rate for 40 and 80 kDa FITC-dextrans. The permeability to carboxyfluorescein was 35 times more than to 80 kDa FITC-dextran [51]. In a study on the permeability of the human ciliary epithelium to a horseradish peroxidase, Tonjum and Pedersen [52] reported that ciliary and iridial epithelium contained a system of paracellular zonulae occludentes. Peroxidase was applied on the stromal side of ciliary body and iris specimens obtained from freshly enucleated eyes. The 40 kDa peroxidase was blocked apically in the lateral intercellular spaces of the CNPE whereas in the iris the progression of peroxidase was blocked apically in the lateral intercellular spaces of the IPE. Freddo [53] studied the intercellular junctions in the posterior IPE cells of the rhesus monkey by electron microscopy, freeze-fracture, and horseradish peroxidase. Intravenously injected horseradish... 

In addition to plant and animal sources, peroxidases are also found in mould, bacteria and microorganisms. A peroxidase from the mould Caldariomyces fumago, chloroperoxidase, has been isolated and characterised. Like the plant peroxidases it has ferriprotoporphyrin IX as the prosthetic group. In many of its chemical and physical properties chloroperoxidase is similar to horseradish peroxidase, but it has the unique ability amongst peroxidases to catalyse the oxidation of chloride ion (Hager et al., 1966 Morris and Hager, 1966). 

However, it should be noted that isoenzymes, i.e., different enzymes catalyzing identical reactions, would have the same four-digit classification. The classification system provides only the basis for a unique identification of an enzyme the particular isoenzyme and its source still have to be specified. For example, peroxidases isolated from soybeans and horseradishes have the same classification, i.e., EC 1.11.1.7. Currently, there are approximately 3200 enzymes that have been listed and assigned classification numbers. 

Aibara S, Yamashita H, Mori E et al (1982) Isolation and characterization of five neutral isoenzymes of horseradish peroxidase. J Biochem 92 531-539... 

Aibara S, Kobayashi T, Morita Y (1981) Isolation and properties of basic isoenzymes of horseradish peroxidase. J Biochem 90 489-496... 

The luciferase isolated from the burrowing clam Pholas dactylus is essentially a peroxidase, which oxidizes a complex oiganic side chain on the surface of a luciferin. This enzyme has not been used in imunoassays however, the luciferin is a substrate for horseradish peroxidase, which produces a very sensitive assay for that enzyme (38). 

Beuzene-1.2-diamine hydrochloride (4.0 g, 22 mmol) was dissolved in distilled Il O (100 mL) in a 250-mL round-bottom flask equipped with a magnetic stirrer. The pH was adjusted to 6.0 with 3 M NaOH. 30% H2O2 (5.0 mL) was then added. The flask was chilled in an ice bath to approximately 10 and horseradish peroxidase (50 mg, 0.001 mol), dissolved in distilled HjO (l.OmL), was added. After 15 min the icc bath was removed and the reaction was allowed to continue for 16 h. The precipitate which had formed was isolated by nitration and rccrystallized from dioxane/benzene yield 2.08 g (42%). 

The covalent binding of N-labelled aniline, in the presence and absence of catalysis by horseradish peroxidase and bimessite, to the fulvic and humic acids isolated from the IHSS Elliot silt loam soil, has been examined by a combination of liquid and solid state N NMR. 

Many of the enzyme/microorganism mediated dealkylation procedures that have been reported have come from the work of Rosazza and his coworkers as a consequence of their studies on Microbial models on Mammalian metabolism. These studies have largely concerned with demethylation of various alkaloids with bacteria and fungi. 0-deethylation reactions with Streptomyces griseus as with dealkylation of 7-ethoxy coumarin gave very poor yields and therefore was not considered a viable synthetic procedure. The quinone imine(I), has been prepared in 64%isolated yield by horseradish peroxidase/hydrogen peroxide O-demethylation of 9-methoxy-ellipticine(II). Studies with H2 0 as a reaction medium demonstrated that the reaction was not a simple demethylation but rather a replacement of the methoxy group by OH from the solvent (as shown in Fig.l)... 

The enzyme most commonly used is horseradish peroxidase (HRP), which is isolated from the root of the horseradish plant. HRP is a 40 kDa enzyme that is bound to antibodies with no loss of either enzyme activity or antibody binding. HRP enzymatic activity is an oxidation reaction that is used for a wide variety of reactions where oxidation gives rise to labeling. 

Horseradish peroxidase. After HRP was treated with 0.1 m l DHF, pH 5.0, the enzyme was isolated by passing the solution through Sephadex G-25 column 51% of HI remained about 40 minutes after the treatment started (solid line). 1 mM dimethyl-p-phemjlenediamine was added to tliis solution at pH 6.0 (broken line). Dotted line is spectrum of the solution in which all of III autodecom-posed to ferriperoxidase... 

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