Separation of horseradish peroxidase isoenzymes by preparative polyacrylamide gel electrophoresis

1977 ◽  
Vol 55 (18) ◽  
pp. 2474-2477
Author(s):  
Sheikh A. Saeed ◽  
Josephine Cuthbert
Keyword(s):  
Horseradish Peroxidase ◽  
Electrophoretic Mobility ◽  
Peroxidase Activity ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Single Step ◽  
Polyacrylamide Gel ◽  
Single Band ◽  
Peroxidase Isoenzymes ◽  
Crude Material

Seven peroxidase isoenzymes were purified from crude horseradish peroxidase (EC 1.11.1.7) (HRP) in a single step by preparative polyacrylamide gel electrophoresis. The peroxidase activity in seven isoenzymes accounted for 90% of the activity in the crude material. Each isoenzyme (except HRP7) after separation migrated as a single band with characteristic electrophoretic mobility. All of the purified isoenzymes were found to retain complete peroxidase activity after storage at −20 °C or 4 °C for 3 months.

scholarly journals Studies on Platelet Glycoproteins by Surface Labeling and SDS Polyacrylamide Gel Electrophoresis

1977 ◽  
Author(s):  
I. Hagen ◽  
N.O. Solum ◽  
M. Peterka
Keyword(s):  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Conformational State ◽  
Polyacrylamide Gel ◽  
The Other ◽  
Metabolic Inhibitors ◽  
Conventional System ◽  
Platelet Surface ◽  
Release Reaction

Platelet surface (glyco)proteins, and alterations in these in connection with the thrombin-induced release reaction has been studied. Platelets were labeled by lactoperoxidase-catalyzed iodination, and examined by SDS gel electrophoresis in two different gel systems, one conventional(J. Biol. Chem.1969 244 4406), and the other containing urea and EDTA in the gels. In the conventional system the bulk of radioactivity coincided with a PAS band (GP III) of MW about 100, 000. In the other system, the main radioactive peak appeared in the GP II area (MW 120,000), and a shift in the PAS stain intensity from GP III to GP II was seen. Thrombasthenic platelets showed only traces of the GP II band in both systems. The bulk of radioactivity was associated with the surface glycopolypeptide GPS, which is present, but not labeled in normal platelets. In thrombin-released platelets, GPS in its unreduced state has an altered electrophoretic mobility compared to control platelets and platelets which have been incubated with metabolic inhibitors to prevent secretion. The findings indicate that the GP III band consists of two different polypeptides, one of which appears in the GP II area in gels containing urea and EDTA. Further, that thrombasthenic platelet membrane exists in a conformational state different from that of normal platelets. And finally, GPS is in some way involved in, or influenced by, the thrombin-induced release reaction.

scholarly journals Purification and characterization of mouse α-lactalbumin and preparation of its antibody

1980 ◽  
Vol 185 (1) ◽  
pp. 227-237 ◽  
Author(s):  
Y Nagamatsu ◽  
T Oka
Keyword(s):  
Concanavalin A ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Deae Cellulose ◽  
Polyacrylamide Gel ◽  
Single Band ◽  
Major Species ◽  
Alpha Lactalbumin

alpha-Lactalbumin was purified to apparent homogeneity from mouse milk by combined use of gel filtration, chromatography on DEAE-cellulose and hydroxyapatite, and concanavalin A-Sepharose affinity chromatography. Mouse alpha-lactalbumin exists in several species with different charges and in two molecular-size forms. The smaller form, which constituted over 90% of total alpha-lactalbumin, included two major and two minor species, each of which showed different electrophoretic mobility on polyacrylamide-gel electrophoresis, but gave the same single band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in two different buffer systems and over the range 10-15% acrylamide concentrations. The molecular weight was estimated as 14 100. The two major species of the smaller form had the same amino acid composition and contained no significant amount of carbohydrate. The larger form of alpha-lactalbumin, consisting of two species with different charges, was present in a small amount (less than 10%) in the milk and was isolated by its ability to interact with concanavalin A-Sepharose. Each of the two species also gave the same single band of apparent mol.w.t 18 500 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. However, this value may be anomalous, since this larger form appears to be glycosylated, and glycoproteins can behave anomalously on sodium dodecyl sulphate/polyacrylamide gels by binding less sodium dodecyl sulphate. All species of mouse alpha-lactalbumin from milk were active in the lactose synthase reaction and showed identical immunological properties, as determined by the mono-specific antibody prepared against the small major species. The presence of both the larger and the smaller forms, each in a percentage concentration similar to that found in milk, was also demonstrated in alpha-lactalbumin induced by hormones in organ cultureof pregnant-mouse mammary gland.

scholarly journals Biology of amazonian mosquitoes. III. Esterase isozymes in Anopheles darlingi.

1985 ◽  
Vol 15 (1-2) ◽  
pp. 167-178 ◽  
Author(s):  
Joselita M.M. dos Santos ◽  
Eucleia P.B. Contel ◽  
Warwick E. Kerr
Keyword(s):  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Developmental Stages ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Anopheles Darlingi ◽  
Genetic Studies ◽  
Esterase Isozymes ◽  
Phenotypic Distribution ◽  
Polymorphic System

Six esterase isozymes were studied during the development of Anopheles darlingi by using polyacrylamide gel electrophoresis and two different substrates, a-naphthylcelate and a-naphthylpropionate. Esterases 5 and 6'were detected in all developmental stages esterases 1 and 2 were more intensively stained if larvae, while esterases 3 and 4 were better visualized in pupae and adults. Strong differences in intensity of some of the isozymes were observed during the pupal stage.Four out of the six isozymes showed variation in the electrophoretic mobility. Esterase-2 was choosed for genetic studies, because was the best stained isozyme in the gels. Two codominant alleles {Est2*S and Est2*F) code for this polymorphic system, with the Est*S frequency equal to 0.521. Phenotypic distribution is in agreement with hardy-Weinberg expectations.

ISOLATION OF CANINE PROLACTIN BY POLYACRYLAMIDE GEL ELECTROPHORESIS

1976 ◽  
Vol 82 (3) ◽  
pp. 691-705 ◽  
Author(s):  
G. E. Jones ◽  
A. D. Brownstone ◽  
A. R. Boyns
Keyword(s):  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Prolactin Secretion ◽  
Disc Electrophoresis ◽  
Polyacrylamide Gel ◽  
Pituitary Extract ◽  
Immunoreactive Material

ABSTRACT The electrophoretic mobility of prolactin obtained from canine pituitary extract was studied with the aid of polyacrylamide disc electrophoresis. Using a preparative gel electrophoretic system the immunoreactive material was purified on a quantitative scale which was then used to develop a homologous radioimmunoassay for canine prolactin. The radioimmunoassay system was able to detect prolactin in the plasma of dogs after the administration of agents which would be expected to affect prolactin secretion.

ROLE OF GALACTOSE IN THE ANTIGENIC PROPERTIES OF THYROGLOBULIN

1976 ◽  
Vol 83 (1) ◽  
pp. 93-98 ◽  
Author(s):  
F. Monaco ◽  
M. Andreoli
Keyword(s):  
Sialic Acid ◽  
Thyroid Nodule ◽  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Human Thyroid ◽  
Antigenic Properties ◽  
Beta Galactosidase

ABSTRACT Thyroglobulins (TG) from a "hot" human thyroid nodule and from Fisher rats have been purified and the effects of progressive removal of sialic acid and galactose on the immunoreactive properties of the proteins were studied. Terminal sialic acid and galactose were released by stepwise hydrolysis with neuraminidase and beta-galactosidase. Agalacto-TG shows a slower electrophoretic mobility than native TG, but in polyacrylamide gel electrophoresis and immunoelectrophoresis it migrates in the same position as asialo-TG. In immunodiffusion agalacto-TG forms a spur with native TG and asialo-TG when tested against anti 19S native TG or anti-asialo-TG sera. It is thus shown that galactose in the terminal environment of the oligosaccharide chains of thyroglobulin is essential for the structural groups involved in the antigenic properties of thyroglobulin.

ELECTROPHORETIC IDENTITY OF BIOLOGICALLY ACTIVE PITUITARY AND SERUM PROLACTINS IN THE LACTATING AND PERPHENAZINETREATED RAT

1973 ◽  
Vol 72 (4) ◽  
pp. 654-662 ◽  
Author(s):  
M. Ben-David ◽  
A. Chrambach
Keyword(s):  
Electrophoretic Mobility ◽  
Gel Electrophoresis ◽  
Post Partum ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Biologically Active ◽  
Neutral Ph

ABSTRACT Polyacrylamide gel electrophoresis at neutral pH, 0°C, was used to test the identity of prolactin derived from serum and pituitary extracts of rats. Animals were studied after treatment with perphenazine, a prolactin releasing agent, and during post partum lactation. After gel electrophoresis, the position on the gel of prolactin activity was determined by the pigeon crop-sac bioassay. The biologically active prolactin exhibited identical electrophoretic mobility, whether derived from serum or pituitary, of either normal lactating or perphenazine treated rats.

scholarly journals A study by polyacrylamide-gel electrophoresis of the effect of proteolysis on Micrococcus lysodeikticus polynucleotide phosphorylase

1968 ◽  
Vol 110 (3) ◽  
pp. 475-479 ◽  
Author(s):  
P. S. Fitt ◽  
E. A. Fitt ◽  
H. Wille
Keyword(s):  
Electrophoretic Mobility ◽  
Degradation Product ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Progressive Increase ◽  
Active Species ◽  
Polyacrylamide Gel ◽  
Polyacrylamide Gels ◽  
Polynucleotide Phosphorylase ◽  
Micrococcus Lysodeikticus

1. Trypsin digestion of Micrococcus lysodeikticus polynucleotide phosphorylase (nucleoside diphosphate–polynucleotide nucleotidyltransferase) causes a progressive increase in electrophoretic mobility in polyacrylamide gels of the single active degradation product. 2. A marked increase in primer requirement for CDP polymerization occurs before a more mobile product is formed. 3. α-Chymotrypsin digestion yields a product that separates into several active species on polyacrylamide-gel electrophoretograms. 4. No separation of ADP-and CDP-polymerization activities occurs during electrophoresis after either trypsin or α-chymotrypsin treatment.

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