scholarly journals Pj variant, a new hybrid MNSs glycoprotein of the human red-cell membrane

1982 ◽  
Vol 203 (2) ◽  
pp. 419-426 ◽  
Author(s):  
D Blanchard ◽  
J P Cartron ◽  
P Rouger ◽  
C Salmon
Keyword(s):  
Sialic Acid ◽  
Blood Group ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Low Frequency ◽  
Periodic Acid Schiff ◽  
Red Cell Membrane ◽  
Sialic Acid Content ◽  
External Domain

An unusual glycoprotein variant (Pj) was found inherited through a caucasian family exhibiting atypical N and Nvg blood-group reactivities. Pj erythrocytes are blood-group-MS homozygous and have a normal sialic acid content. On sodium dodecyl sulphate/polyacrylamide-gel electrophoresis the variant contains a new component Pj of 24kDa apparent molecular mass in the monomeric state which is sharply stained by periodic acid/Schiff reagent. Both blood-group-MN (alpha) and -Ss (delta) glycoproteins were present. Homodimers (Pj2) as well as heterodimers with MN-glycoprotein (alpha Pj) and the Ss-glycoprotein (delta Pj) were also identified. The new sialoglycoprotein Pj is trypsin- and chymotrypsin-resistant in situ and carries N- and Nvg- but not M- and S-reactivities. The Pj component is labelled by lactoperoxidase-catalysed radioiodination. A 3H label is also easily introduced into the sialic acid or the galactose and galactosamine of the Pj glycoprotein. It is proposed that the Pj is a hybrid glycoprotein containing the N-terminal end of delta-glycoprotein and the C-terminal end of the alpha-glycoprotein. This proposal is supported by the finding that Pj carries a leucine residue at its N-terminus and is not immunoprecipitated by a monoclonal mouse antibody (R18) reacting specifically with the external domain of glycoprotein alpha. The red cells from the proposita Pj were found positive for a very low frequency MN antigen named Sta.

scholarly journals Platelet membrane studies in the May-Hegglin anomaly

Blood ◽  
1981 ◽  
Vol 58 (2) ◽  
pp. 279-284 ◽  
Author(s):  
BS Coller ◽  
MH Zarrabi
Keyword(s):  
Sialic Acid ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Glycoprotein ◽  
Galactose Oxidase ◽  
Periodic Acid Schiff ◽  
Giant Platelets ◽  
Total Sialic Acid ◽  
Bernard Soulier Syndrome

Abstract Since studies of the giant platelets in the Bernard-Soulier syndrome have shown decreased electrophoretic mobility, decreased sialic acid, and an abnormality in a membrane glycoprotein, we performed similar studies on the giant platelets from two patients with the May-Hegglin anomaly. The patients' platelet electrophoretic mobilities did not differ from control. Although the total sialic acid contents of the patients' platelets were greater than control when calculated per platelet, they were very similar to control when normalized for differences in platelet volume and surface area. When platelet proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis there were no differences between the glycoproteins of control and patient platelets as judged by the patterns of periodic acid Schiff staining and fluorescein-labeled concanavalin A binding. Similarly, patterns of surface glycoprotein labeling by neuraminidase/galactose oxidase/KB3H4 were identical. We conclude that unlike the giant platelets in the Bernard-Soulier syndrome, those of the May-Hegglin anomaly are not associated with a membrane abnormality detectable by these techniques.

scholarly journals Miltenberger Class I and II erythrocytes carry a variant of glycophorin A

1983 ◽  
Vol 213 (2) ◽  
pp. 399-404 ◽  
Author(s):  
D Blanchard ◽  
A Asseraf ◽  
M J Prigent ◽  
J P Cartron
Keyword(s):  
Blood Group ◽  
Rare Variants ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Class I ◽  
Glycophorin A ◽  
M Gene ◽  
Periodic Acid Schiff ◽  
Unusual Variant

The membranes from Miltenberger Class I (Mi I) and II (Mi II) erythrocytes, two rare variants at the blood group MNSs locus, exhibited an abnormal glycoprotein of 32 kDa apparent molecular mass sharply stained by the periodic acid/Schiff procedure and a decreased content of glycoprotein alpha (synonym glycophorin A, glycoprotein MN) as seen on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Purified 125I-labelled Vicia graminea lectin binds to the unusual 32 kDa glycoprotein separated from Mi I and Mi II erythrocyte membrane of blood group NN or MN, but no significant labelling of this band was observed with Mi samples typed MM. On the basis of such lectin-labelling experiments we have described two heterozygous MN, Mi I individuals that carry one copy of an M gene producing a normal alpha-glycoprotein with M-specificity and one copy of a MiI gene producing a 32 kDa glycoprotein with N-specificity. Further investigations have shown that the 32 kDa glycoprotein was immunoprecipitated by two mouse monoclonal antibodies (R18 and R10) reacting specifically with the external domain of glycoprotein alpha. These results demonstrate that Mi I and Mi II erythrocytes carry an unusual variant of glycoprotein alpha.

Factor VIII/Von Willebrand Factor Protein Sensitivity Of Periodic Acid Schiff Stain To Carbohydrate Deficiency

1981 ◽  
Author(s):  
H Gralnick ◽  
G Jackson ◽  
S Williams ◽  
E Cregger
Keyword(s):  
Sialic Acid ◽  
Von Willebrand Factor ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Staining Reaction ◽  
Periodic Acid Schiff ◽  
Sialic Acid Content ◽  
Pas Staining ◽  
Von Willebrand ◽  
Willebrand Factor

Carbohydrate (C) deficiency of the factor VUI/von Willebrand factor (f. VIII/vWf) protein has been implicated as one of the molecular defects of von Willebrand’s disease. The periodic acid Schiff (PAS) reaction in polyacrylamide gel electrophoresis (PAGE) has been used as a screening test for C deficiency. To ascertain the sensitivity of this reaction to the C content of the f. VIII/vWf protein we have purified the normal protein and compared the Coomassie to PAS staining reaction of the subunit of the intact, asialo (neuraminidase-treated), asialo-agalacto (neuraminidase and β-galactosidase-treated) derivatives and a fourth derivative which was treated with a mixed glycosidase which included neuraminidase, β-galactosidase and N-acetyl-glucosaminidase. Gels were run simultaneously, one stained with Coomassie blue and the other with PAS and at various time intervals after staining the gels were scanned.Studies of the intact f. VIII/vWf protein revealed that the PAS and Coomassie stains could be compared from 24 hours up to 7 days with no significant changes. The desialated f. VIII/vWf protein had insignificant differences in the PAS stain compared to normal (range 81-108% of intact). However, the asialo-agalacto derivative (36% of intact) and the f. VIII/vWf treated with the mixed glycosidase (24% of intact) had significantly reduced PAS staining in relation to normal. C analysis of the different f. VIII/vWf proteins revealed that removal of the sialic acid reduced the C 30% while in the asialo-agalacto the C was reduced 47% and in the mixed glycosidase-treated material the C was reduced 78%. The PAS stain is not very sensitive to the sialic acid content of the f. VIII/vWf protein and reduction of 30% of the C does not affect the PAS stain. Since the PAS stain is relatively insensitive to significant carbohydrate deficiencies of the f. VIII/vWf protein, the use of the PAS stain in PAGE may not be an adequate test to ascertain C content of the f. VIII/vWf protein.

scholarly journals Platelet membrane studies in the May-Hegglin anomaly

Blood ◽  
1981 ◽  
Vol 58 (2) ◽  
pp. 279-284 ◽  
Author(s):  
BS Coller ◽  
MH Zarrabi
Keyword(s):  
Sialic Acid ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Glycoprotein ◽  
Galactose Oxidase ◽  
Periodic Acid Schiff ◽  
Giant Platelets ◽  
Total Sialic Acid ◽  
Bernard Soulier Syndrome

Since studies of the giant platelets in the Bernard-Soulier syndrome have shown decreased electrophoretic mobility, decreased sialic acid, and an abnormality in a membrane glycoprotein, we performed similar studies on the giant platelets from two patients with the May-Hegglin anomaly. The patients' platelet electrophoretic mobilities did not differ from control. Although the total sialic acid contents of the patients' platelets were greater than control when calculated per platelet, they were very similar to control when normalized for differences in platelet volume and surface area. When platelet proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis there were no differences between the glycoproteins of control and patient platelets as judged by the patterns of periodic acid Schiff staining and fluorescein-labeled concanavalin A binding. Similarly, patterns of surface glycoprotein labeling by neuraminidase/galactose oxidase/KB3H4 were identical. We conclude that unlike the giant platelets in the Bernard-Soulier syndrome, those of the May-Hegglin anomaly are not associated with a membrane abnormality detectable by these techniques.

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.

scholarly journals Impact of Botrytis cinerea Contamination on the Characteristics and Foamability of Yeast Macromolecules Released during the Alcoholic Fermentation of a Model Grape Juice

Molecules ◽  
2020 ◽  
Vol 25 (3) ◽  
pp. 472
Author(s):  
Richard Marchal ◽  
Thomas Salmon ◽  
Ramon Gonzalez ◽  
Belinda Kemp ◽  
Céline Vrigneau ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Alcoholic Fermentation ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Grape Juice ◽  
Size Exclusion ◽  
Periodic Acid Schiff ◽  
Exclusion Chromatography ◽  
The Impact

Botrytis cinerea is a fungal pathogen responsible for the decrease in foamability of sparkling wines. The proteolysis of must proteins originating from botrytized grapes is well known, but far less information is available concerning the effect of grape juice contamination by Botrytis. The impact from Botrytis on the biochemical and physico-chemical characteristics of proteins released from Saccharomyces during alcoholic fermentation remains elusive. To address this lack of knowledge, a model grape juice was inoculated with three enological yeasts with or without the Botrytis culture supernatant. Size exclusion chromatography coupled to multi-angle light scattering (SEC-MALLS) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) techniques (AgNO3 and periodic acid Schiff staining) was used in the study. When Botrytis enzymes were present, a significant degradation of the higher and medium MW molecules released by Saccharomyces was observed during alcoholic fermentation whilst the lower MW fraction increased. For the three yeast strains studied, the results clearly showed a strong decrease in the wine foamability when synthetic musts were inoculated with 5% (v/v) of Botrytis culture due to fungus proteases.

scholarly journals Detection in milk of a 60,000 Mr mouse and a 68,000 Mr human phosphorylated glycoprotein by histochemical analysis on polyacrylamide gels.

1983 ◽  
Vol 31 (6) ◽  
pp. 709-716 ◽  
Author(s):  
M R Green
Keyword(s):  
Molecular Weight ◽  
Sialic Acid ◽  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Blue Staining ◽  
Polyacrylamide Gels ◽  
Periodic Acid Schiff ◽  
Coomassie Blue Staining ◽  
Coomassie Blue

Proteins in colostrum and skimmed milk from humans and mice were separated by electrophoresis on polyacrylamide gels and stained with Coomassie blue (CB), Ethyl-Stains-all (ESA), and periodic acid-Schiff (PAS) to investigate changes that may occur in milks throughout lactation. In mouse colostrum but not in mature mouse milk, a PAS-positive protein of apparent molecular weight of 60,000 stained prominently blue with ESA. A protein in human milk with a molecular weight of 68,000 stained similarly but was present throughout lactation. The intensity of blue staining of these minor proteins in milk approached that obtained with casein phosphoproteins. The metachromatic dye ESA stains phosphoproteins and sialic acid-rich glycoproteins blue to blue-green. Removal of phosphorus from the former and sialic acid from the latter results in those proteins staining red with ESA. The intensity of blue staining of the 60,000 and 68,000 Mr proteins was diminished but not lost following treatment with phosphatase. It was eliminated following neuraminidase digestion of the mouse protein and mild acid hydrolysis of the human protein. Coomassie blue staining of the proteins was not affected by these procedures. Following electrophoresis of milk and milk fractions in a non-sodium dodecyl sulfate-containing system, the proteins were identified by their characteristic staining properties with ESA and isolated.

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.

Cell surface characteristics of Mortierella species and their interaction with a mycoparasite

1984 ◽  
Vol 30 (3) ◽  
pp. 290-298 ◽  
Author(s):  
M. S. Manocha
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Characteristics ◽  
Protein Patterns ◽  
Periodic Acid Schiff ◽  
The Difference ◽  
Chitin And Chitosan

Cell surface characteristics of three Mortierella species differing in their response to a mycoparasite, Piptocephalis virginiana, were examined. Their cell wall composition was typical of mucoraceous fungi with chitin and chitosan as major polysaccharides. Electron microscopy revealed that the mycoparasite penetrated and formed haustoria in the hyphae of susceptible hosts, M. pusilla and M. isabellina. The failure of the parasite to establish contact and penetrate a hypha of the nonhost, M. candelabrum, was not due to cell wall thickness, rigidity, or chitin contents. Markedly different protein patterns obtained from crude alkali extracts of host and nonhost cell walls by sodium dodecyl sulfate – polyacrylamide gel electrophoresis might explain the difference in host and nonhost response to the mycoparasite. Whereas most of the bands differed only in intensity after staining with either Coomassie blue or periodic acid – Schiff reagent, there were two distinct bands of glycoproteins (76 000 and 74 000) observed in the host species which were absent in the nonhost species.

Studies of the heterogeneity of human pituitary LH by fast protein liquid chromatography

1985 ◽  
Vol 105 (3) ◽  
pp. 405-413 ◽  
Author(s):  
A. Stockell Hartree ◽  
J. B. Lester ◽  
R. C. Shownkeen
Keyword(s):  
Liquid Chromatography ◽  
Sialic Acid ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Binding Activity ◽  
Fast Protein Liquid Chromatography ◽  
Human Pituitary ◽  
Sialic Acid Content ◽  
Receptor Binding Activity

ABSTRACT The Pharmacia fast protein liquid chromatography system was employed to fractionate a purified preparation of human LH (hLH) on the anion exchanger Mono Q at pH 7·8 into 14 sub-fractions. Each of the sub-fractions was characterized by its behaviour on polyacrylamide gel electrophoresis, sodium dodecyl sulphate (SDS) gel electrophoresis, LH receptor binding activity and sialic acid content. All sub-fractions contained sialic acid, were active in binding to LH receptors, and exhibited components typical of hLH subunits on SDS gel electrophoresis. None of the sub-fractions was homogeneous with respect to charge. There is evidence that part of the heterogeneity results from the presence in some molecules of an internal proteolytic cleavage within the β-subunit, and fractions enriched in species containing such cleavages were prepared by this method. J. Endocr. (1985) 105,405–413

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