scholarly journals The purification and properties of the lectin from potato tubers, a hydroxyproline-containing glycoprotein

1973 ◽  
Vol 135 (2) ◽  
pp. 307-314 ◽  
Author(s):  
Anthony K. Allen ◽  
Albert Neuberger
Keyword(s):  
Amino Acid ◽  
Wheat Germ ◽  
Gel Filtration ◽  
Indirect Evidence ◽  
Basic Amino Acid ◽  
Potato Tubers ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Sepharose 4B ◽  
Abundant Amino Acid

1. Potato lectin has been purified and shown to be a glycoprotein containing about 50% of carbohydrate. Most of the sugar residues (92%) are arabinose; small amounts of galactose, glucose and glucosamine are also present. 2. The most abundant amino acid is hydroxyproline (16% of the residues), 11.5% of the residues are half-cystine and phenylalanine is absent. The lectin also contains about one residue/molecule of a basic amino acid, not usually found in proteins, which has been tentatively identified as ornithine. There is indirect evidence that the components of the glycoprotein are linked through hydroxyproline and arabinose. 3. By gel filtration in 6m-guanidine–HCl on Sepharose 4B, it was found that both the native glycoprotein and its S-carboxymethylated derivative had subunit molecular weights of 46000 (±5000). In a non-denaturing solution, two of these units appear to be associated. 4. The lectin is specifically inhibited in its agglutination reaction by oligosaccharides that contain N-acetylglucosamine. Its specificity is similar to, but not identical with, that of wheat-germ agglutinin.

scholarly journals Purification and properties of α-galactosidases from Vicia faba seeds

1969 ◽  
Vol 113 (1) ◽  
pp. 49-55 ◽  
Author(s):  
P. M. Dey ◽  
J. B. Pridham
Keyword(s):  
Amino Acid ◽  
Vicia Faba ◽  
Aromatic Amino Acid ◽  
Gel Filtration ◽  
Disc Electrophoresis ◽  
Thermal Stabilities ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Enzyme I ◽  
Carbohydrate Contents

Two forms of α-galactosidase, I and II, exist in Vicia faba seeds and these have been purified 3660- and 337-fold respectively. They behaved as homogeneous preparations when examined by ultracentrifugation, disc electrophoresis and gel filtration. The apparent molecular weights of enzymes I and II, as determined by gel filtration, were 209000 and 38000 respectively. The carbohydrate contents of enzymes I and II were 25% and 2·8% respectively, and the enzymes differed in their aromatic amino acid compositions. Enzyme I was split into six inactive subunits in the presence of 6m-urea. α-Galactosidases I and II showed different pH optima and Km and Vmax. values with p-nitrophenyl α-d-galactoside and raffinose as substrates, and also differed in their thermal stabilities.

scholarly journals The purification and properties of the l-serine O-sulphate-degrading system of pig liver

1971 ◽  
Vol 121 (5) ◽  
pp. 747-752 ◽  
Author(s):  
N. Tudball ◽  
P. Thomas ◽  
R. Bailey-Wood
Keyword(s):  
Amino Acid ◽  
Isoelectric Focusing ◽  
Enzyme System ◽  
Gel Filtration ◽  
Pig Liver ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Isoelectric Points ◽  
Degrading System ◽  
Similar Amino Acid

1. The enzyme system from pig liver responsible for the αβ-elimination of l-serine O-sulphate was purified 1000-fold. 2. Isoelectric focusing produced two enzymically active fractions with isoelectric points at pH5.6 and 5.9 respectively. 3. Osmometry and gel filtration showed both enzymes to possess molecular weights of approx. 54000. 4. The separate activities exhibited similar amino acid compositions.

Effect of extractant and molecular size on the optical and chemical properties of soil humic acids

Soil Research ◽  
1969 ◽  
Vol 7 (3) ◽  
pp. 229 ◽  
Author(s):  
JHA Butler ◽  
JN Ladd
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Humic Acids ◽  
Gel Filtration ◽  
Chemical Properties ◽  
Molecular Size ◽  
Carboxyl Content ◽  
Peptide Bonds ◽  
Molecular Weights ◽  
Amino Acid Contents

Humic acids extracted from soil with sodium pyrophosphate have greater proportions of lower molecular weight material, less acid-hydrolysable amino acid nitrogen contents, but greater carboxyl contents and extinction values (260 and 450 nm) than humic acids extracted subsequently from the same sample with alkali. Humic acids extracted with alkali from fresh soil samples have intermediate values. Extinction values at 260 nm are directly correlated with carboxyl contents for a given soil. Different crop histories have no significant effect on the measured properties of the extracted humic acids. An alkali-extracted humic acid has been fractionated by gel filtration into seven fractions of different nominal molecular weight ranges. As the molecular weights of the fractions increase, both aliphatic C-H (based on infrared absorption at 2900 cm-1) and acid-hydrolysable amino acid contents increase, whereas extinction values at 260 nm and carboxyl contents decrease. The infrared spectra of the high molecular weight fractions have peaks at 1650 and 1510 cm-1 which correlate with acid-hydrolysable amino acid contents and which correspond to amide I and II bands of peptide bonds. Alkaline hydrolysis to split peptide bonds eliminates both these peaks. The spectra also have peaks at 1720 and 1210 cm-1 which correlate with the carboxyl content.

scholarly journals α-Crystallin. The isolation and characterization of distinct macromolecular fractions

1971 ◽  
Vol 124 (2) ◽  
pp. 337-343 ◽  
Author(s):  
Abraham Spector ◽  
Lu-Ku Li ◽  
Robert C. Augusteyn ◽  
Arthur Schneider ◽  
Thomas Freund
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Chemical Difference ◽  
Molecular Weights ◽  
Isolation And Characterization ◽  
Different Populations ◽  
Molecular Weight Fractions

α-Crystallin was isolated from calf lens periphery by chromatography on DEAE-cellulose and gel filtration. Three distinct populations of macromolecules have been isolated with molecular weights in the ranges approx. 6×105−9×105, 0.9×106−4×106and greater than 10×106. The concentration of macromolecules at the molecular-weight limits of a population are very low. The members of the different populations do not appear to be in equilibrium with each other. Further, in those molecular-weight fractions investigated, no equilibrium between members of the same population was observed. The population of lowest molecular weight comprises 65–75% of the total material. The amino acid and subunit composition of the different-sized fractions appear very similar, if not identical. The only chemical difference observed between the fractions is the presence of significant amounts of sugar in the higher-molecular-weight fractions. Subunit molecular weights of approx. 19.5×103and 22.5×103were observed for all α-crystallin fractions.

scholarly journals HUMAN AMYLOID PROTEIN: CHEMICAL VARIABILITY AND HOMOGENEITY

1971 ◽  
Vol 19 (1) ◽  
pp. 1-15 ◽  
Author(s):  
M. HARADA ◽  
C. ISERSKY ◽  
P. CUATRECASAS ◽  
D. PAGE ◽  
H. A. BLADEN ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amyloid Fibril ◽  
Gel Filtration ◽  
Protein S ◽  
Amyloid Protein ◽  
X Ray Diffraction ◽  
Amyloid Proteins ◽  
X Ray ◽  
Chemical Variability ◽  
Sepharose 4B

The morphology of the fibril of amyloid derived from different individuals is similar, but occasionally significant differences are noted. All human amyloid filaments have a "β-pleated sheet" conformation as revealed by x-ray diffraction, and those examined after orientation show a "cross-β" pattern. All amyloid fibril concentrates studied so far can be fractionated to obtain the major amyloid protein component(s) by sequential gel filtration with 5 M guanidine-HCl in 1 N acetic acid on Sepharose 4B and Sephadex G-100 or G-75 columns with the removal of over 28% of proteins representing minor constituents. The major amyloid protein(s) obtained from the spleen and/or liver of six patients is found to contain tryptophan, to be deficient in hydroxylysine and hydroxyproline and usually at least one commonly occurring amino acid and to have a high content of dicarboxylic acid and short chain amino acids and unreactive (blocked) NH2-terminal groups or aspartic acid-asparagine (Asx). However, the amyloid protein(s) from each individual differs from that of the others in molecular weight, in amino acid composition and in the presence or absence of specific tryptic peptides. Amyloid protein(s) from the liver and spleen of the same individual is identical. No chemical characteristics distinguish amyloid proteins derived from cases classified clinically as "primary" from those classified as "secondary." There is a striking chemical similarity between amyloid proteins and the NH2-terminal variable fragment of the light and heavy chain of immumoglobulin proteins.

Cellular Amino Acid Concentrations and Regulation of Aspartate Kinase in the Thermophilic Phototrophic Prokaryote Chloroflexus aurantiacus

1990 ◽  
Vol 45 (1-2) ◽  
pp. 74-78 ◽  
Author(s):  
Jobst-Heinrich Klemme ◽  
Gisela Laakmann-Ditges ◽  
Jutta Mertschuweit
Keyword(s):  
Amino Acid ◽  
Feedback Inhibition ◽  
Gel Filtration ◽  
Break Point ◽  
Test System ◽  
Chloroflexus Aurantiacus ◽  
Aspartate Kinase ◽  
Homoserine Dehydrogenase ◽  
Molecular Weights ◽  
Amino Acid Concentrations

Aspartate kinase (AK , EC 2.7.2.4) from the thermophilic, phototrophic prokaryote, Chloroflexus aurantiacus, was partially purified and separated from homoserine dehydrogenase (HSDH, EC 1.1.1.3). The molecular weights as determined by gel filtration were 130,000 and 46,000, respectively. HSDH had a moderately high thermal stability (50% inactivation at 84 °C) and displayed its activity optimum at 72 °C. By contrast, AK had its activity optimum at 52 °C (with a break-point in the Arrhenius plot at 42 °C) and was much less thermostable (50% inactivation at 67 °C). The Km-values for aspartate and ATP (determined in a pyruvate kinase-coupled test system) were 10.5 and 0.63 mM , respectively. The enzyme was strongly inhibited by L-threonine (Ki = 10 μm) and activated by alanine, isoleucine, valine and methionine. L-Threonine acted as a mixed-type inhibitor in respect to aspartate, and non-competitively in respect to ATP. Contrary to AKs from Rhodospirillaceae, the enzyme from Chloroflexus aurantiacus was not subject to a concerted feedback inhibition by two amino acids of the aspartate family. The regulatory properties of the aspartate kinase are discussed in relation to the cellular amino acid concentrations.

scholarly journals Preparation of high Fischer ratio oligopeptide by proteolysis of corn gluten meal

2008 ◽  
Vol 26 (No. 1) ◽  
pp. 38-47 ◽  
Author(s):  
Ma Ying ◽  
Lin Li ◽  
Sun Da-Wen
Keyword(s):  
Amino Acid ◽  
Gel Filtration ◽  
Aromatic Amino Acids ◽  
Enzyme Reaction ◽  
Ion Exchange Resin ◽  
Orthogonal Experimental Design ◽  
Degree Of Hydrolysis ◽  
Corn Gluten Meal ◽  
Molecular Weights ◽  
Corn Gluten

A method to obtain an oligopeptide with high Fischer ratio is described. Corn gluten meal (CGM) was hydrolysed with Alcalase 2.4L using a two-step hydrolysis. In the first-step hydrolysis, the enzyme reaction conditions for hydrolysing CGM were optimised by using the orthogonal experimental design, while pH = 8.0, temperature = 55°C, enzyme to substrate ratio (3:97, w/w), and the substrate concentration = 5% were identified as the optimum conditions, under which up to 11.62% degree of hydrolysis (DH) could be obtained. The hydrolysate was then fractionated by ultrafiltration using a membrane with the molecular cutoff of over 10 kD at 20 kPa. For the second-step hydrolysis, the filtrate was adjusted to pH 6.0, then papain was added at 50°C and the mixture was maintained for 3 hours. The hydrolysate was obtained after inactivating papain and centrifuging. Then the salt (mainly NaCl) in the hydrolysate was removed with an ion exchange resin at the speed of 8 times bed volume per hour, and aromatic amino acids were removed through absorption by active carbon. By using Sephadex G-25 gel filtration chromatography, a peptide mixture with low molecular weights between 1000 and 1300 was obtained. Finally, tests on amino acid composition and free amino acid concentration of oligopeptide solution showed that the oligopeptide had a high Fischer ratio of 34.71 and the yield of 11.59%.

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.

scholarly journals Carcinoembryonic antigen-like substances of human urothelial carcinomas. Isolation of components from pathological urine and comparison with colorectal carcinoma antigens

1974 ◽  
Vol 139 (2) ◽  
pp. 431-440 ◽  
Author(s):  
R. Nery ◽  
A. L. Barsoum ◽  
H. Bullman ◽  
A. M. Neville
Keyword(s):  
Carcinoembryonic Antigen ◽  
Gel Filtration ◽  
Molecular Size ◽  
Density Gradient Ultracentrifugation ◽  
Colorectal Carcinomas ◽  
Net Charge ◽  
Molecular Weights ◽  
Urothelial Carcinomas ◽  
Sepharose 4B ◽  
Mean Molecular Weight

Urines of several patients with urothelial carcinomas contain inhibitors of the immunoreaction between carcinoembryonic antigen derived from human colorectal carcinomas and monospecific goat antiserum raised against the antigen. These inhibitors range in approximate molecular weights from less than 1000 to several millions, and two have been isolated by a combination of extraction, gel filtration and electrophoretic procedures. These are respectively a macromolecular aggregate, component UCEA-3, which is excluded by Sepharose 4B, and a glycoprotein(s) component, UCEA-1, with mean molecular weight (2×105) similar to that of carcinoembryonic antigen. Comparison of the properties of component UCEA-1 and carcinoembryonic antigen on gel filtration, electrophoresis, immunoelectrophoresis and density gradient ultracentrifugation indicates that these substances of similar molecular size and net charge differ in some immunochemical properties.

scholarly journals The isolation of three neurophysins from porcine posterior pituitary lobes

1970 ◽  
Vol 116 (5) ◽  
pp. 899-909 ◽  
Author(s):  
L. O. Uttenthal ◽  
D. B. Hope
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Analysis ◽  
Gel Filtration ◽  
Biological Significance ◽  
Starch Gel Electrophoresis ◽  
Posterior Pituitary ◽  
Fresh Tissue ◽  
Molecular Weights ◽  
Molecular Dimensions

1. Three neurophysins, proteins that bind the polypeptide hormones oxytocin and vasopressin, have been isolated from acetone-dried porcine posterior pituitary lobes. The proteins have been named porcine neurophysins-I, -II and -III in order of their electrophoretic mobilities at pH8.1. 2. Electrophoretic comparison of the purified proteins, which are homogeneous on starch-gel electrophoresis, with the soluble proteins of fresh porcine posterior pituitary lobes extracted in 0.1m-HCl and in buffer pH8.1 suggests that the isolated proteins are native to the fresh tissue. 3. Neurophysins-I and -II are present in similar amounts in the tissue, whereas neurophysin-III is present only in small quantities. Acetone-dried tissue also contains traces of other hormone-binding neurophysin components. 4. All the neurophysins can bind both oxytocin and [8-lysine]-vasopressin. 5. The apparent molecular weights of the neurophysins increase with increasing protein concentration as measured by equilibrium sedimentation in the ultracentrifuge. 6. Neurophysins-I and -III are of similar molecular dimensions, contain one residue of methionine per molecule and lack histidine. The minimum molecular weight of neurophysin-I obtained by amino acid analysis is 9360. Neurophysin-II is of larger molecular dimensions than neurophysins-I and -III and can be separated from these by gel filtration on Sephadex G-75. It contains no histidine or methionine, and its minimum molecular weight has been estimated as 14020 by amino acid analysis. 7. Each of the three neurophysins possesses N-terminal alanine. 8. The possible biological significance of the existence of several neurophysins within one species is discussed.

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