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.

Purification and Properties of γ-Glutamyl Cyclotransferase from Pig Liver

1971 ◽  
Vol 49 (2) ◽  
pp. 218-226 ◽  
Author(s):  
E. D. Adamson ◽  
A. Szewczuk ◽  
G. E. Connell
Keyword(s):  
Amino Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ammonium Sulfate Precipitation ◽  
Cyclic Derivative ◽  
Pig Liver ◽  
Molecular Weights ◽  
Purification And Properties ◽  
Isoelectric Points ◽  
Pyrrolidone Carboxylic Acid ◽  
Amino Acid Glycine

The widely occurring enzyme γ-glutamyl cyclotransferase acts on γ-glutamyl peptides to effect the release of the terminal glutamyl residue as the cyclic derivative pyrrolidone carboxylic acid. The enzyme has been purified from pig liver by (1) ammonium sulfate precipitation from the supernatants of homogenates, (2) CM-cellulose treatment, (3) DEAE-Sephadex chromatography, and (4) preparative polyacrylamide gel electrophoresis. The homogeneity of the highly purified enzyme was demonstrated by ultracentrifugation and electrophoretic analyses. By means of (5) isoelectric focusing, two forms of the enzyme with isoelectric points 4.87 and 4.95 were separated. These forms proved to have very similar sedimentation velocities, molecular weights, and amino acid compositions, and to have the same amino acid, glycine, as the N-terminal residue.

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 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.

A comparison of four sulfhydryl cathepsins (B, C, H, and L) from porcine spleen

1984 ◽  
Vol 62 (12) ◽  
pp. 1301-1308 ◽  
Author(s):  
K. R. Lynn ◽  
Rosalind S. Labow
Keyword(s):  
Molecular Weight ◽  
High Pressure ◽  
Cathepsin L ◽  
Gel Filtration ◽  
Cathepsin C ◽  
Molecular Weights ◽  
Isoelectric Points ◽  
Dipeptidyl Aminopeptidase ◽  
Precise Assessment ◽  
Similar Amino Acid

Four sulfhydryl cathepsins, B, C (dipeptidyl aminopeptidase I), H, and L were isolated from porcine spleen. They are all glycoproteins of similar amino acid compositions, which are comparable with those of cathepsins B and H from other sources and so with papain. All four cathepsins exist in multiple charged forms: B, C, H, and L have isoelectric points in the range 4.3–5.4, 5.3 and 5.9, 5.2–5.7, and 7–8.7, respectively. The molecular weights of cathepsins B and H were 24 000 and 26 000. Anomalous behaviour of cathepsin L on both conventional gel filtration and high pressure liquid chromatography precluded a precise assessment of its weight which is between 22 000 and 28 000. The isolated mercurial derivative of cathepsin C has a molecular weight of 56 000 (an active dimer formed on reduction). Cathepsins B and H also aggregate.

scholarly journals The Proteins of the Keratin Component of Bird?s Beaks

1976 ◽  
Vol 29 (6) ◽  
pp. 467 ◽  
Author(s):  
MJ Frenkel ◽  
JM Gillespie
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Major Protein ◽  
Major Fraction ◽  
Amino Acid Compositions ◽  
Molecular Weights ◽  
Isoelectric Points ◽  
Related Proteins ◽  
Keratin Proteins ◽  
Similar Amino Acid

Birds' beaks have an outer shell of hard keratin which consists almost entirely of proteins which are very rich in glycine [about 30 residues per 100 residues (residues %)], contain moderate levels of tyrosine and serine (each about 8 residues %), and which have relatively low contents of cystine (about 2�5 residues %), lysine, histidine, isoleucine and methionine. Major protein fractions in the S-carboxymethyl form isolated from the beaks of 'six different orders of birds have similar amino acid compositions, isoelectric points (pH 4�2-4�9) and molecular weights (13 000--14 500). Detailed chromatographic electrophoretic and compositional studies of the proteins of kookaburra beak reveal them to be a family of closely related proteins with only limited heterogeneity, in contrast to mammalian keratin systems. The major kookaburra beak fraction is similar in overall composition and molecular weight to fowl epidermal scale, kookaburra claw and turtle scute proteins and shows some resemblance to reptile claw protein. Beaks also contain small amounts of protein which are distinctly different from the major fraction but which resemble feather keratin proteins in composition and size.

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.

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%.

scholarly journals Purification and properties of a new testosterone 17β-dehydrogenase (NADP+) from guinea-pig liver

1977 ◽  
Vol 163 (3) ◽  
pp. 401-407 ◽  
Author(s):  
E Kaguera ◽  
S Toki
Keyword(s):  
Guinea Pig ◽  
Column Chromatography ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Supernatant Fraction ◽  
Pig Liver ◽  
Ring Junction ◽  
Purification And Properties ◽  
Androstane Derivatives ◽  
Guinea Pig Liver

As a result of studies of guinea-pig live testosterone 17beta-dehydrogenase (NADP+) (EC 1.1.1.64), a new testosterone 17beta-dehydrogenase was discovered. The new enzyme was purified to a single homogeneous protein from the 105 000 g-supernatant fraction of guinea-pig liver by (NH4)2SO4 fractional precipitation and two gel-filtration stages, DEAE-cellulose column chromatography and hydroxyapatite column chromatography. It was characterized by many properties. The enzyme has almost the same properties as the classical testosterone 17beta-dehydrogenase (NADP+) (EC 1.1.1.64), with respect to cofactor requirement, pH optima for dehydrogenation, effect of phosphate ion on the NAD+-dependent reaction and molecular weight, but characteristic differences were observed in substrate-specificity between the two dehydrogenases. With various androstane derivatives, the configuration of the A/B-ring junction was closely connected with enzyme activity. 5alpha-Androstanes, such as 5alpha-androstane-3alpha,17beta-diol, 5alpha-androstane-3beta,17beta-diol and 17beta-hydroxy-5alpha-androstan-3-one, and 5beta-congeners, such as 5beta-androstane-3alpha,17beta-diol, 5beta-androstane-3beta,17beta-diol and 17beta-hydroxy-5beta-androstan-3-one, served as substrates for both the EC 1.1.1.64 enzyme and the new enzyme. The EC 1.1.1.64 enzyme oxidized testosterone more rapidly than did the new enzyme. These comparisons were based on the relative activities, apparent Km values and apparent Vmax values.

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