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.

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 Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

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 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 MURINE AMYLOID FIBRIL PROTEIN: ISOLATION, PURIFICATION AND CHARACTERIZATION

1971 ◽  
Vol 19 (1) ◽  
pp. 16-28 ◽  
Author(s):  
G. G. GLENNER ◽  
D. PAGE ◽  
C. ISERSKY ◽  
M. HARADA ◽  
P. CUATRECASAS ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Disc Electrophoresis ◽  
Structure Identification ◽  
Major Protein ◽  
Amyloid Protein ◽  
Molecular Weights ◽  
Sepharose 4B

Murine amyloid has been produced by four different induction methods ( Mycobacterium butyricum, casein, casein plus Freund's adjuvant and endotoxin-induced mouse amyloidosis) in several strains and obtained from mice with "spontaneous" amyloidosis. The amyloid fibrils have been concentrated from spleen and liver. Electron microscopy of all of these preparations reveals the amyloid fibril to be 100 Å in width and composed of two parallel filaments, each measuring 35-40 Å in width and having the appearance of a twisted ribbon. X-ray diffraction of all preparations reveals a "backbone" spacing at 4.75 Å and a "side chain" spacing at 11 Å indicating a "β-pleated sheet" structure. Identification of the major protein component of amyloid fibril concentrates was made by combined use of sodium dodecyl sulfate polyacrylamide disc electrophoresis, labeling of the protein with 3H-tryptophan and Sepharose 4B gel filtration. Purification of the amyloid protein from spontaneous amyloidosis liver was accomplished by sequential gel filtration with 5 M guanidine in 1 N acetic acid on Sepharose 4B and Sephadex G-100 or G-75 columns. The material is a unique glycoprotein with a molecular weight of 7200, a high content of dicarboxylic and short chain amino acids, a significant amount of tryptophan and an unreactive NH2-terminal amino acid, tentatively identified as pyrrolid-2-one-5-carboxylic acid. There are no methionine, half-cystine, hydroxylysine or hydroxyproline residues. Murine amyloid protein, therefore, has striking similarities to many human amyloid protein preparations. It differs from the human proteins in the similarity of molecular weights of different preparations.

Determination Of Molecular Size Of VIII:C In Whole Plasma By Electron Irradiation

1981 ◽  
Author(s):  
J Harmon ◽  
G A Jamieson ◽  
G Rock
Keyword(s):  
Molecular Weight ◽  
Electron Irradiation ◽  
Gel Filtration ◽  
Molecular Size ◽  
Target Size ◽  
Platelet Poor Plasma ◽  
Molecular Weights ◽  
Normal Plasma ◽  
Biological Mixtures

Loss of activity during electron irradiation provides a means of determining the molecular size of specific molecules in complex biological mixtures. This technique has established a molecular weight of 200,000 for Factor VIII:C in concentrates prepared from citrated plasma in which calcium is sequestered by chelation (Aronson, et al. Thromb. Diath. Haemorrh. 8:270, 1962): this value is similar to that obtained by more conventional techniques. However recent data suggest that in heparinized plasma, where physiological levels of calcium are maintained, about half of the VIII:C activity has a molecular weight of 50,000 as determined by gel filtration and ultracentrifugation (Rock, et al. Thromb. Res. 13:85, 1978). When heparinized plasma was subjected to electron irradiation in the frozen state there was a perceptible loss of VIII:C activity at 1 megarad and 80% loss with 60 megarads of irradiation. Analysis of the course of inactivation showed a biphasic curve with 73% of the VIII:C activity having a target size of 40,000 daltons while 28% had a molecular weight in excess of one million. Similar results were obtained when blood was collected in citrate and rapidly processed (∼5 min) to platelet-poor plasma. Following electron irradiation, 62% of VIII:C activity showed a target size of 35,000 daltons while the remaining 38% gave a target size of 275,000. These results provide further evidence that circulating VIII:C activity in normal plasma has a molecular weight of about 40,000 and suggest that reports of higher molecular weights are an artifact of the chelation of calcium as evidenced by the biphasic decay of VIII:C activity in citrated plasma.

Interaction Of Atherogenic (VLdL; LdL) And Non Atherogenic Lipoproteins (HDL) With Heparin Fractions Of Various Molecular Size

1981 ◽  
Author(s):  
P Duthilleul ◽  
P Fievet ◽  
L Bouillet ◽  
J C Fruchart
Keyword(s):  
Gel Filtration ◽  
Molecular Size ◽  
Two Dimensional ◽  
Fluorimetric Method ◽  
Molecular Weights ◽  
Crossed Immunoelectrophoresis ◽  
At Iii ◽  
Heparin Fractions ◽  
Inhibition Of Thrombin

Some anticoagulant properties of various heparin fractions (with molecular weights from 5 000 to 20 300 daltons) have been investigated in purified and plasmatic systems with and without Lipoproteins isolated by ultracentrifugation from hyperlipemic subjects (type IIa). Amidolytic (Tos-Gly-Pro-Arg-pNA) and coagulometric (Yin and Wessler) anti Xa activity of a HMW fraction (231 USP/mg. MW: 20 300) was decreased from 15 %m both systems supplied with LDL or VLDL since HDL have no effect. No influence was detected on anti Ila activity (Phe-pro-arg-AIE fluorimetric method). LMW (20 USP/mg ; MW 6 000 - 8 000) and ULMW (20 USP.MW 4 000-5 000) anti Xa and anti Ila activities was modified neither by VLDL/LDL neither by HDL. These findings point out to differences in the mechanisms of inhibition of thrombin and Xa by various molecular weigh heparin fractions.Two dimensional crossed Immunoelectrophoresis in agarose 1 % with mixing various quantities of each heparin fraction (0 - 100 μg/ml) in the first phase of electrophoresis of LDL/VLDL, LDL/VLDL and AT III, LDL/VLDL and FXa reveal the presence of LDL or VLDL/heparin fractions complexes and LDL or VLDL/AT III complexes, which are isolated by Gel filtration on Ultrogel A 6. We conclude that FXa can escape from neutralization by inactive complexes.

Suppressive Effect of Dextran on Platelet Adhesiveness

1966 ◽  
Vol 16 (03/04) ◽  
pp. 384-394 ◽  
Author(s):  
S Cronberg ◽  
B Robertson ◽  
Inga Marie Nilsson ◽  
J.-E Niléhn
Keyword(s):  
Molecular Weight ◽  
Bleeding Time ◽  
Slight Modification ◽  
Molecular Size ◽  
Suppressive Effect ◽  
Factor Ix ◽  
Factor V ◽  
Platelet Adhesiveness ◽  
History Of ◽  
Mean Molecular Weight

Summary43 normal volunteers, 3 patients with thrombophlebitis, and 1 patient with a high platelet adhesiveness and a history of thrombophlebitis have received dextran and its action on the mechanism of haemostasis has been studied. Platelet adhesiveness has been investigated by a slight modification of Hellem’s methods for whole blood and plasma. Dextran with a mean molecular weight of 70,000 produced a markedly lowered platelet adhesiveness together with a moderate prolongation of the Ivy bleeding time. Factor VIII was decreased by about 50% and factor V, factor IX and fibrinogen were decreased slightly more than could be expected from haemodilution alone. No fibrinolysis occurred. Dextran of lower molecular size was less potent. The possible use of dextrans as a thrombosis prophylactic agent is discussed.

scholarly journals Purification and characterization of fat body lipase from the greater wax moth, Galleria mellonella (Lepidoptera: Pyralidae)

2019 ◽  
Vol 81 (1) ◽  
Author(s):  
Rahma R. Z. Mahdy ◽  
Shaimaa A. Mo’men ◽  
Marah M. Abd El-Bar ◽  
Emad M. S. Barakat
Keyword(s):  
Metal Ions ◽  
Kinetic Parameters ◽  
Biochemical Characterization ◽  
Galleria Mellonella ◽  
Fat Body ◽  
Specific Activity ◽  
Larval Instar ◽  
Gel Filtration ◽  
Molecular Weights

Abstract Background Insect lipid mobilization and transport are currently under research, especially lipases and lipophorin because of their roles in the production of energy and lipid transport at a flying activity. The present study has been conducted to purify intracellular fat body lipase for the first time, from the last larval instar of Galleria mellonella. Results Purification methods by combination of ammonium sulfate [(NH4)2SO4] precipitation and gel filtration using Sephadex G-100 demonstrated that the amount of protein and the specific activity of fat body lipase were 0.008633 ± 0.000551 mg/ml and 1.5754 ± 0.1042 μmol/min/mg protein, respectively, with a 98.9 fold purity and recovery of 50.81%. Hence, the sephadex G-100 step was more effective in the purification process. SDS-PAGE and zymogram revealed that fat body lipase showed two monomers with molecular weights of 178.8 and 62.6 kDa. Furthermore, biochemical characterization of fat body lipase was carried out through testing its activities against several factors, such as different temperatures, pH ranges, metal ions, and inhibitors ending by determination of their kinetic parameters with the use of p-nitrophenyl butyrate (PNPB) as a substrate. The highest activities of enzyme were determined at the temperature ranges of 35–37 °C and 37–40 °C and pH ranges of 7–9 and 7–10. The partially purified enzyme showed significant stimulation by Ca2+, K+, and Na+ metal ions indicating that fat body lipase is metalloproteinase. Lipase activity was strongly inhibited by some inhibitors; phenylmethylsulfonyl fluoride (PMSF), ethylene-diaminetetractic acid (EDTA), and ethylene glycoltetraacetic acid (EGTA) providing evidence of the presence of serine residue and activation of enzymes by metal ions. Kinetic parameters were 0.316 Umg− 1 Vmax and 301.95 mM Km. Conclusion Considering the purification of fat body lipase from larvae and the usage of some inhibitors especially ion chelating agents, it is suggested to develop a successful control of Galleria mellonella in near future by using lipase inhibitors.

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