The Properties of Soluble Proteins Synthesized by Nephrotic Rat Kidney Microsomes

1972 ◽  
Vol 50 (3) ◽  
pp. 292-298
Author(s):  
V. N. Katiyar ◽  
B. L. Dinh
Keyword(s):  
Gel Electrophoresis ◽  
Rat Kidney ◽  
Periodic Acid ◽  
Rabbit Antiserum ◽  
Periodic Acid Schiff ◽  
Rat Serum ◽  
Tissue Protein ◽  
Schiff Reagent ◽  
Normal Rat ◽  
Kidney Microsomes

The incorporation of 14C-leucine into the microsomal proteins of nephrotic rat kidney was much higher than that in the microsomal proteins of the normal rat kidney. When the newly synthesized microsomal proteins were analyzed by acrylamide gel electrophoresis, it was found that the higher incorporation of 14C-leucine in nephrotic rat kidney was mostly due to an increase in the biosynthesis of a tissue protein which migrated in the electrophoretic zone between serum albumin and transferrin. This protein did not react with rabbit antiserum to normal rat serum and was stained with periodic acid – Schiff reagent. It was believed to be excreted in the urine of nephrotic rats in great quantity and was easily identified as a distinct band in electrophoregrams of urine from these rats.

scholarly journals The Structural Characteristics of Chicken Fibrinogen

1977 ◽  
Author(s):  
J. Pindyck ◽  
M. W. Mosesson ◽  
D. Bannerjee ◽  
D. Galanakis
Keyword(s):  
Gel Electrophoresis ◽  
Structural Characteristics ◽  
Periodic Acid ◽  
Factor Xiiia ◽  
Periodic Acid Schiff ◽  
Polymer Formation ◽  
Sensitive Site ◽  
Schiff Reagent ◽  
The One ◽  
Fibrinogen Molecule

The structure and subunit composition of chicken fibrinogen(ϕ) have been investigated. Dodecyl sulfate gel electrophoresis of unreduced specimens revealed a single ϕ band with a molecular weight of approximately 320,000. ϕ and fibrin specimens were also electrophoresed after reduction with dithiothreitol, and after crosslinking of unreduced specimens in the presence of Factor Xllla. Chromatographically separated S-sulfo chains were also studied after reptilase or thrombin treatment,and certain samples were stained with periodic acid Schiff reagent(PAS). Chicken Aα chains weresmaller than human Aα chains (54,500 vs.70,900, respectively) but, like mammalian Aα chains, they possessed a reptilase and thrombin sensitive site, were PAS negative,and undergo Factor XIIIa catalyzed α-polymer formation. The sizes of chicken Bβ and γ chains were nearly thesame as their mammalian counterparts, (i. e. 60,000 and 49,000 respectively) ; both types of chains were PAS positive. Chicken Bβ chains possessed a slowly reactive thrombin sensitive site apparently corresponding to the one in human ϕ; the chicken β chains, like mammalian β chains, did not undergo Factor XIIIa catalyzed cross-linking. Like mammalian γ chains, chicken γ chains could undergo Factor XIIIa catalyzed γ-γ dimerization and did not possess thrombin or reptilase sensitive sites. These findings indicate that the chicken fibrinogen molecule is composed of three pairs of disulfide-bridged chains corresponding in most respects to mammalian fibrinogen chains.

scholarly journals Purification of the major endoglucanase from Aspergillus fumigatus Fresenius

1983 ◽  
Vol 213 (2) ◽  
pp. 437-444 ◽  
Author(s):  
J B Parry ◽  
J C Stewart ◽  
J Heptinstall
Keyword(s):  
Aspergillus Fumigatus ◽  
Isoelectric Focusing ◽  
Gel Electrophoresis ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Periodic Acid Schiff ◽  
Schiff Reagent ◽  
Solubilizing Activity

Aspergillus fumigatus (Fresenius), IMI 246651, A.T.C.C. 46324, produces two β-glucosidase enzymes, cotton-solubilizing activity, xylanase and endoglucanase enzymes which can be separated by gel-filtration chromatography. The major endoglucanase does not bind to concanavalin A-Sepharose and does not stain with periodic acid/Schiff reagent. It is homogeneous on polyacrylamide isoelectric focusing (pI = 7.1) and has a mol.wt. of 12500 by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The endoglucanase produces glucose and a mixture of oligosaccharides from cellulose; the purified enzyme has a small dextranase activity. It is stable at 50 degrees C and pH 6.

scholarly journals Purification, characterization and radioimmunoassay of adenosine deaminase from human leukaemic granulocytes

1981 ◽  
Vol 195 (2) ◽  
pp. 389-397 ◽  
Author(s):  
D A Wiginton ◽  
M S Coleman ◽  
J J Hutton
Keyword(s):  
Molecular Weight ◽  
Adenosine Deaminase ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Periodic Acid Schiff ◽  
Apparent Homogeneity

Adenosine deaminase was purified 3038-fold to apparent homogeneity from human leukaemic granulocytes by adenosine affinity chromatography. The purified enzyme has a specific activity of 486 mumol/min per mg of protein at 35 degrees C. It exhibits a single band when subjected to sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, non-denaturing polyacrylamide-gel electrophoresis and isoelectric focusing. The pI is 4.4. The enzyme is a monomeric protein of molecular weight 44000. Both electrophoretic behaviour and molecular weight differ from those of the low-molecular-weight adenosine deaminase purified from human erythrocytes. Its amino acid composition is reported. Tests with periodic acid-Schiff reagent for associated carbohydrate are negative. Of the large group of physiological compounds tested as potential effectors, none has a significant effect. The enzyme is specific for adenosine and deoxyadenosine, with Km values of 48 microM and 34 microM respectively. There are no significant differences in enzyme function on the two substrates. erythro-9-(2-Hydroxy non-3-yl) adenine is a competitive inhibitor, with Ki 15 nM. Deoxycoformycin inhibits deamination of both adenosine and deoxyadenosine, with an apparent Ki of 60-90 pM. A specific antibody was developed against the purified enzyme, and a sensitive radioimmunoassay for adenosine deaminase protein is described.

Quantitative scanning of glycoproteins on polyacrylamide gels stained with periodic acid-schiff reagent (PAS)

1973 ◽  
Vol 55 (1) ◽  
pp. 313-316 ◽  
Author(s):  
Jean-Marie Matthieu ◽  
Richard Hudson Quarles
Keyword(s):  
Periodic Acid ◽  
Polyacrylamide Gels ◽  
Periodic Acid Schiff ◽  
Schiff Reagent

Molecular weight determination and partial characterization of Klebsiella pneumoniae hemolysins

1986 ◽  
Vol 32 (11) ◽  
pp. 884-888 ◽  
Author(s):  
Lucila Isabel Barberis ◽  
Alberto Jorge Eraso ◽  
Maria Cristina Pàjaro ◽  
Inès Albesa
Keyword(s):  
Klebsiella Pneumoniae ◽  
Gel Electrophoresis ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Polyacrylamide Gel ◽  
Growth Media ◽  
Molecular Weight Determination ◽  
Periodic Acid Schiff

Two thiol-activated Klebsiella pneumoniae hemolysins were purified from growth media by means of salt precipitation, gel filtration, ion-exchange chromatography, and polyacrylamide gel electrophoresis. The hemolysins peaks coincided with the protein and glycoprotein peaks as determined by chromatography and electrophoresis, The molecular weights, estimated by gel filtration, were 8400 and 19 000; by sodium dodecyl sulphate–polyacrylamide gel electrophoresis, the values were calculated as 15 500 and 27 000. The electrophoretic bands were best detected by the periodic acid–Schiff method. Reduction of the disulfide linkages did not cause the originally larger molecule to break into 8400 and 19 000 hemolysins. However, trypsin treatment cleaved the 19 000 hemolysin into an active moiety, with an electrophoretic migration similar to the 8400 hemolysin. A naturally occurring proteolytic activity was investigated using pepstatin and antipain. When the trypsin inhibitor was added to the system, the hemolytic activity was detected only in the 19 000 hemolysin and the smaller hemolysin was absent.

scholarly journals IMMUNOLOGIC STUDIES OF HEART TISSUE

1961 ◽  
Vol 113 (1) ◽  
pp. 1-16 ◽  
Author(s):  
Melvin H. Kaplan ◽  
Frederick D. Dallenbach
Keyword(s):  
Heart Disease ◽  
Connective Tissue ◽  
Gamma Globulin ◽  
Fluorescent Antibody ◽  
Periodic Acid ◽  
Heart Tissue ◽  
Periodic Acid Schiff ◽  
Schiff Reagent ◽  
Points Of View ◽  
Rheumatic Heart

Using fluorescent antibody methods, deposits of bound gamma globulin, as determined in unfixed washed sections of auricular appendages from rheumatic hearts, were noted in a significant number (18 per cent) of 100 specimens studied. Such deposits were observed in myofibers, sarcolemma, interstitial connective tissue, and vessel walls. Albumin and fibrin were generally found absent from these sites. Control hearts from normal and pathologic material, including postmortem and biopsied specimens, in general, did not reveal such deposits. These various tissue sites which contained bound gamma globulin frequently exhibited evidence of alteration as indicated both by enhanced affinity for eosin and by strongly positive reaction with the periodic acid-Schiff reagent, and appeared comparable in some cases to "fibrinoid." Bound gamma globulin was not observed in cellular or stromal components of Aschoff lesions, nor was the occurrence of Aschoff lesions correlated with presence of bound gamma globulin. It is suggested that deposition of gamma globulin and the eosinophilic alteration associated with such deposition are related to certain of the pathologic changes of rheumatic heart disease. The nature of such deposits of gamma globulin was considered from immune and non-immune points of view.

Isolation and partial characterization of a soluble mucin in human pancreatic juice

1991 ◽  
Vol 69 (8) ◽  
pp. 566-571
Author(s):  
S. P. Lee ◽  
Y. S. Choong ◽  
H. Z. Park
Keyword(s):  
Molecular Weight ◽  
Pancreatic Juice ◽  
Periodic Acid ◽  
Average Molecular Weight ◽  
Small Molecular Weight ◽  
Periodic Acid Schiff ◽  
Cellulose Acetate Electrophoresis ◽  
Cscl Density Gradient ◽  
Oligosaccharide Moiety ◽  
Schiff Reagent

Morphological and histochemical abnormalities in pancreatic mucin occur in many pancreatic disorders. However, the composition of pancreatic mucin is poorly understood. Purified mucin was isolated from pure pancreatic juice by sequential chromatography on Sepharose CL-2B and CL-4B followed by CsCl density gradient ultracentrifugation. The mucin preparation consists of 24% protein and 73% carbohydrate. Reduction of the macromolecule (> 2 × 106) by mercaptoethanol resulted in the formation of subunits of molecular weight 500 000 and released several small molecular weight proteins, including a glycoprotein of an average molecular weight of 116 000. Cellulose acetate electrophoresis separated the mucin into three species of different staining properties for periodic acid-Schiff reagent and Alcian blue, suggesting the presence of microheterogeneity with respect to sulphation and sialation. Threonine, serine, and proline composed 48% of the total amino acids, while the oligosaccharide moiety contained N-acetylglucosamine, N-acetyl-galactosamine, fucose, galactose, sialic acid, and sulphate. We also detected the presence of C16:0 and C18:0 fatty acids which were probably noncovalently bound to the pancreatic mucin.Key words: pancreas, mucin, glycoprotein, cystic fibrosis.

Human intestinal goblet cell mucin

1976 ◽  
Vol 54 (8) ◽  
pp. 707-716 ◽  
Author(s):  
I. Jabbal ◽  
D. I. C. Kells ◽  
G. Forstner ◽  
J. Forstner
Keyword(s):  
Goblet Cell ◽  
High Speed ◽  
Proteolytic Enzymes ◽  
Periodic Acid ◽  
Periodic Acid Schiff ◽  
Strong Concentration ◽  
Acid Ratio ◽  
Schiff Reagent ◽  
Major Chemical ◽  
Sepharose 4B

Goblet cell mucin (GCM) has been purified for the first time from mucosal scrapings of human small intestine. Proteolytic enzymes and organic solvents were avoided during the isolation procedure. The mucin was purified by Sepharose 4B and 2B column chromatography of high-speed supernatant fractions. The most purified fraction was compared with rat intestinal GCM. The two were similar with respect to chemical composition, antigenic features, and polyacrylamide disc gel electrophoresis. The major chemical differences included a higher hexosamine–fucose and hexosamine – sialic acid ratio in human mucin. The two mucins showed strong concentration dependence in sedimentation velocity studies. Human mucin at a concentration of 0.2 to 1.5 mg protein per millilitre gave multiple associated peaks with variable s0 values (10.8–36.6). Rat mucin, in contrast, gave a constant (although polydisperse) pattern with s0 = 15.15. To explore these differences both mucins were stained with periodic acid – Schiff reagent and subjected to band ultracentrifugation at concentrations of 0.6–1.9 μg protein per millilitre. At this low concentration, rat mucin did not change in its sedimentation characteristics. In contrast, human GCM produced a single peak with s0 = 37.9. Thus dilution abolished polydispersity in the human but not the rat mucin, suggesting that intermolecular bonding forces in the human mucin are weaker.

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