Cytoplasmic Carboxylesterases of Human and Domestic Animal Liver: Aggregation, Dissociation and Molecular Weight Estimation

1972 ◽  
Vol 50 (1) ◽  
pp. 9-15 ◽  
Author(s):  
D. J. Ecobichon
Keyword(s):  
Molecular Weight ◽  
Human Liver ◽  
Gel Filtration ◽  
Domestic Animal ◽  
Starch Gel Electrophoresis ◽  
Weight Estimation ◽  
Animal Liver ◽  
Starch Gel ◽  
Species Specific ◽  
Two Peaks

The cytoplasmic carboxylesterases of bovine, ovine, equine and human liver were fractionated by starch gel electrophoresis and by gel filtration on Sephadex. While species-specific, heterogeneous bands were observed in starch gel, the esterases of the bovine, ovine and equine liver were eluted from Sephadex G-100 as single peaks of activity, each with a characteristic elution volume. Gel filtration of human liver extracts yielded two peaks of activity, one containing electrophoretically slow esterases, the other electrophoretically fast esterases. Extracted equine and human hepatic carboxylesterases aggregated readily on storage or concentration, forming larger units which could be dissociated by a combination of acidic pH and high salt concentration. Molecular weight estimates of the hepatic esterases by gel filtration on Sephadex G-100 and G-200 yielded values of 65 000 for ovine, 55 000 for bovine, 96 000 and 70 000 for equine variants and 180 000 and 65 000 for human variants. The observations suggested that the cytoplasmic enzymes in relatively crude hepatic extracts had a lower molecular weight than those in concentrated or partially purified preparations which formed stable dimers or trimers.

Bovine hepatic carboxylesterases chromatographic fractionation, gel filtration, and molecular weight estimation

1969 ◽  
Vol 47 (8) ◽  
pp. 799-805 ◽  
Author(s):  
D. J. Ecobichon
Keyword(s):  
Molecular Weight ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Bovine Liver ◽  
Weight Estimation ◽  
Molecular Weights ◽  
Starch Gel ◽  
Two Peaks ◽  
Second Peak

The carboxylesterase activity of bovine liver was fractionated by DEAE-cellulose chromatography into two peaks of activity. Electrophoresis in starch gel showed that the peak eluted first was composed of a group of five electrophoretically slow bands while the second peak was a single, rapidly migrating band. Gel filtration of the crude extract on Sephadex G-100 and G-200 yielded a single peak of esterase activity containing both the electrophoretically slow and fast bands. The determination of molecular weights by gel filtration on Sephadex G-200 and G-100 yielded estimates of 52 000 and 55 000, respectively. The molecular weight estimates of the DEAE-cellulose fractionated electrophoretically slow and fast bands on Sephadex G-100 were identical, namely 55 000.

scholarly journals A chromatographic preparation of ox growth hormone

1966 ◽  
Vol 100 (3) ◽  
pp. 593-600 ◽  
Author(s):  
M Wallis ◽  
HBF Dixon
Keyword(s):  
Molecular Weight ◽  
Growth Hormone ◽  
Gel Electrophoresis ◽  
Anterior Pituitary ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Starch Gel Electrophoresis ◽  
Highly Active ◽  
Cross Linkage ◽  
Starch Gel

1. A method is described for the chromatographic preparation of ox growth hormone. It involves chromatography of an extract of anterior pituitary lobes on DEAE-cellulose, followed by rechromatography on a dextran gel of low cross-linkage (Sephadex G-100). 2. The product is highly active in growth-hormone assays, and is obtained in good yield. It was homogeneous by several criteria, but showed some heterogeneity on starch-gel electrophoresis. 3. The molecular weight of the hormone was estimated from its behaviour on gel-filtration columns under various conditions. Evidence that the hormone may dissociate into sub-units under some conditions is presented.

Purification and properties of crustacyanin

1966 ◽  
Vol 164 (994) ◽  
pp. 130-151 ◽  
Keyword(s):  
Molecular Weight ◽  
Quaternary Structure ◽  
Gel Filtration ◽  
Potassium Thiocyanate ◽  
Starch Gel Electrophoresis ◽  
Prosthetic Group ◽  
Reversible Dissociation ◽  
Purification And Properties ◽  
Starch Gel ◽  
Quaternary Structures

Crustacyanin, the blue carapace pigment of the common lobster Homarus gammarus (L.), has been purified and crystallized. This chromoprotein has a minimum molecular weight of 36 000 based on the content of the carotenoid prosthetic group astaxanthin. The molecular weight in gel filtration measurements is about 650 000, corresponding to some 18 molecules of astaxanthin per molecule of protein. Crustacyanin, on dialysis against water, dissociates into particles of about 35 000 molecular weight, each apparently bearing one molecule of carotenoid. The dissociation is accompanied by a shift in the principal maximum of the absorption spectrum from 633 to 595 nm and is reversed upon addition of salt. Reversible dissociation also occurs in the presence of 3 M urea, 1 M potassium thiocyanate, 10% (v/v) dioxan or 10% (v/v) acetone. When the carotenoid is removed from crustacyanin with acetone, the resul­tant apoprotein has a mean molecular weight of about 20 000. It may be resolved by starch gel electrophoresis into several components of which two predominate. Crustacyanin, indistinguishable from the native material, can be reconstituted from apoprotein and carotenoid. Evidence from the behaviour of crustacyanin and its apoprotein at surfaces indicates that the tertiary and quaternary structures of the native protein are stabilized by the carotenoid. It is suggested that the quaternary structure of crustacyanin is induced by an interaction of the carotenoid molecules of the subunits, which in turn causes a change in configuration of the protein favourable to aggregation. The result is a micelle-like structure with a hydrophobic carotenoid core.

scholarly journals The separation and characterization of marmoset kidney β-d-galactosidase and β-d-glucosidase

1977 ◽  
Vol 167 (3) ◽  
pp. 765-773 ◽  
Author(s):  
R J Pierce ◽  
R G Price
Keyword(s):  
Molecular Weight ◽  
Gel Electrophoresis ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Apparent Molecular Weight ◽  
Exchange Chromatography ◽  
Starch Gel Electrophoresis ◽  
Lysosomal Fraction ◽  
Glucosidase Activity ◽  
Starch Gel

beta-D-Galactosidase and beta-D-glucosidase activities were determined in homogenates of marmoset kidney by using the appropriate 4-methylumbelliferyl glycoside, beta-D-Galactosidase activity was separated into two main components by ion-exchange chromatography on DEAE-cellulose, starch-gel electrophoresis, isoelectric focusing and gel filtration on Sephadex G-200. One form designated A had a pI of 5.1, was loosely bound to DEAE-cellulose at pH7.0, remained near the origin on starch-gel electrophoresis at pH 7.0 and had an apparent molecular weight of 160000. The second beta-D-galactosidase component, designated B, was associated with the total beta-D-glucosidase activity, had a pI of 4.3, was firmly bound to DEAE-cellulose, migrated rapidly towards the anode on starch-gel electrophoresis and had an apparent molecular weight of 50000. The optimum pH values of beta-D-galactosidase A and B were 4.5 and 6.0 respectively. beta-D-Galactosidase A was activated by 0.1 M-NaC1 but the activity of the B form was inhibited by 1 M-NaC1 at pH 4.5. beta-D-galactosidase had a bimodal distribution, the A form being recovered in the lysosomal fraction whereas the B form was present in the soluble fraction, as was the major portion of the beta-D-glucosidase activity. The lysosomal and soluble forms were further characterized by DEAE-cellulose chromatography.

Enzyme variation in the Anopheles gambiae Giles group of species (Diptera: Culicidae)

1978 ◽  
Vol 68 (1) ◽  
pp. 85-97 ◽  
Author(s):  
S. J. Miles
Keyword(s):  
Anopheles Gambiae ◽  
Natural Populations ◽  
Lactic Dehydrogenase ◽  
Starch Gel Electrophoresis ◽  
Malic Dehydrogenase ◽  
Glycerophosphate Dehydrogenase ◽  
Starch Gel ◽  
Anopheles Gambiae Giles ◽  
Species Specific ◽  
Glucose 6 Phosphate Dehydrogenase

AbstractThe genotypes of chromosomally-identified individuals from natural populations of the known species of the group of Anopheles gambiae Giles were scored for the enzyme protein structural loci coding for adenylate kinase (Adk), α-naphthyl acetate esterase (Est-1, Est-2, Est-3), glutamic-oxaloacetic transaminase (Got), α-glycerophosphate dehydrogenase (αGpd), hexokinase (Hk), isocitric dehydrogenase (Idh), lactic dehydrogenase (Ldh), ‘leucine’ aminopeptidase (Lap-2), malic enzyme (Me), octanol dehydrogenase (Odh), phosphoglucomutase (Pgm-1, Pgm-2), 6-phosphogluconic dehydrogenase (6-Pgd), phosphohexose isomerase (Phi) and superoxide dismutase (Sod), following starch gel electrophoresis. In the material examined, Est-1, Est-2, Est-3, Got, ldh, Lap-2, Odh, Pgm-1, Pgm-2 and Sod were segregating for two or more alleles; unique alleles at the Est-1, Got and Sod loci produced species-specific phenotypes in A. melas (Theo.), species C and species D, respectively. The further sampling of A. merus Dön, populations supported the presence of a unique SOD phenotype by which this species can also be identified. Of the other enzyme systems examined, no activity following electrophoresis was detected for aldolase and fructose-1,6-diphosphatase, and the resolution of acid and alkaline phosphatase, glyceraldehyde-3-phosphate dehydrogenase, glucose-6-phosphate dehydrogenase, malic dehydrogenase and xanthine dehydrogenase was too poor under the particular electrophoretic conditions for genetic analyses of the enzyme phenotypes.

IDENTIFICATION AND PURIFICATION OF A NEW PROTEIN FROM WATER-SOLUBLE EXTRACTS OF HUMAN ADRENAL GLAND

1979 ◽  
Vol 82 (3) ◽  
pp. 383-NP ◽  
Author(s):  
M. A. AL-AWQATI ◽  
Y. B. GORDON ◽  
T. CHARD
Keyword(s):  
Molecular Weight ◽  
Adrenal Gland ◽  
Gel Filtration ◽  
Water Soluble ◽  
Double Immunodiffusion ◽  
Molecular Weights ◽  
Physicochemical Methods ◽  
Two Peaks ◽  
Sepharose 4B ◽  
New Protein

An homogenate of human foetal adrenal gland was subjected to negative immunoabsorption by column chromatography using anti-whole human serum coupled to Sepharose 4B. Two peaks were eluted and used to immunize rabbits. The antisera produced were absorbed and tested for specificity by double immunodiffusion. Two antigens, which appeared to be specific to the adrenal gland, were identified having molecular weights of 25 000 and 65 000 as determined by gel filtration. The lower molecular weight antigen was isolated by physicochemical methods and found to be a protein. The amino acid composition is reported.

FAMILIAL GOITRE WITH ABSENCE OF THYROGLOBULIN AND SYNTHESIS OF THYROID HORMONES FROM THYROIDAL ALBUMIN

1974 ◽  
Vol 60 (3) ◽  
pp. 389-397 ◽  
Author(s):  
K. B. DESAI ◽  
M. N. MEHTA ◽  
M. C. PATEL ◽  
S. M. SHARMA ◽  
L. RAMANNA ◽  
...  
Keyword(s):  
Thyroid Hormones ◽  
Radioactive Iodine ◽  
Gradient Centrifugation ◽  
Gel Filtration ◽  
Sucrose Density Gradient ◽  
Starch Gel Electrophoresis ◽  
Sucrose Density Gradient Centrifugation ◽  
Salting Out ◽  
Immunological Tests ◽  
Starch Gel

SUMMARY Two siblings, a brother (H. B.) and a sister (R. B.) with long standing goitres were investigated. Radioactive iodine uptake by the thyroid was increased and a significant portion of the plasma radioactive iodine was not extractable with butanol. Chromatography of butanol extracts of serum after radioactive iodine administration showed distinct peaks of triiodothyronine and thyroxine. Microscopic examination of the surgical specimens of the goitres showed Hürthle cell carcinoma with follicles devoid of colloid in both specimens. Sucrose density gradient centrifugation, gel filtration on Sephadex G-200, salting out procedures, starch gel electrophoresis and immunological tests of the supernatant soluble fraction of thyroid homogenates showed a lack of thyroglobulin. Further fractionation of the soluble proteins showed that albumin was apparently involved in the synthesis of thyroid hormones in the absence of thyroglobulin.

scholarly journals Chromatography of transforming deoxyribonucleic acid on methylated albumin and by gel filtration

1974 ◽  
Vol 137 (3) ◽  
pp. 543-546
Author(s):  
G. R. Barker ◽  
P. Hodges
Keyword(s):  
Molecular Weight ◽  
Bacillus Subtilis ◽  
High Molecular Weight ◽  
Deoxyribonucleic Acid ◽  
Degradation Products ◽  
Gel Filtration ◽  
Agarose Gel ◽  
Transforming Activity ◽  
Different Strains ◽  
Two Peaks

1. Native DNA from two strains of Bacillus subtilis was chromatographed by stepwise elution from MAK (methylated albumin on kieselguhr). 2. Transforming activity was confined to two out of the three main fractions, activity being distributed between the two peaks differently for DNA from the different strains. 3. Fractionation of DNA from both strains on 2% agarose gel gave two components. Approx. 75% of the material was eluted within the void volume of the column. Approx. 25% of the material consisted of degradation products of lower molecular weight. 4. Chromatography on MAK of the material of high molecular weight eluted from agarose gel gave a number of peaks differing in molecular weight, indicating that degradation of the DNA takes place during chromatography on MAK. 5. The distribution of transforming activity among the fractions from MAK suggests that degradation occurs preferentially in certain regions of the DNA.

scholarly journals Organic pyrophosphates as substrates for human alkaline phosphatases

1967 ◽  
Vol 105 (3) ◽  
pp. 1307-1312 ◽  
Author(s):  
R. Helen Eaton ◽  
D W Moss
Keyword(s):  
Gel Electrophoresis ◽  
Human Liver ◽  
Inorganic Phosphate ◽  
Starch Gel Electrophoresis ◽  
Alkaline Phosphatases ◽  
Starch Gel ◽  
Pyrophosphatase Activity ◽  
Small Intestinal ◽  
Hydrolysis Of ◽  
Single Enzyme

1. Purified human liver and small-intestinal alkaline orthophosphatases release inorganic phosphate at appreciable rates from a variety of organic pyrophosphate substrates. 2. The pyrophosphatase action is inhibited by Mg2+ ions at concentrations that activate the hydrolysis of orthophosphate substrates by these enzymes. 3. The results of mixed-substrate experiments, denaturation studies with heat or urea and starch-gel electrophoresis suggest that both orthophosphatase and pyrophosphatase activities are, in each preparation, properties of a single enzyme. 4. Intestinal phosphatase shows greater pyrophosphatase activity relative to orthophosphatase than the liver enzyme.

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