[5] Molecular weight determination of glycoproteins by polyacrylamide gel electrophoresis in sodium dodecyl sulfate

Segrest, Jere P.; Jackson, Richard L.

doi:10.1016/0076-6879(72)28007-7

Cited by 763 publications

(310 citation statements)

References 7 publications

Supporting

Mentioning

292

Contrasting

Unclassified

Order By: Relevance

“…6 a). Desialylation of glycoproteins can result in either a decrease (Doms et al, 1989) or increase (Segrest & Jackson, 1972) in electrophoretic mobility. Presumably, this reflects an interplay between effects on shape versus charge: the removal of sialic acid would facilitate molecular sieving, whereas the loss of its negative charge would reduce the negative charge of the protein unless compensated by increased binding of SDS.…”

Section: G Protein Synthesized In the Presence Of Bfa Contains Sialicmentioning

confidence: 99%

Oligomerization and post-translational processing of glycoprotein G of human respiratory syncytial virus: altered O-glycosylation in the presence of brefeldin A

Collins

Mottet²

1992

Journal of General Virology

103

View full text Add to dashboard Cite

Section: G Protein Synthesized In the Presence Of Bfa Contains Sialicmentioning

confidence: 99%

Oligomerization and post-translational processing of glycoprotein G of human respiratory syncytial virus: altered O-glycosylation in the presence of brefeldin A

Collins

Mottet²

1992

Journal of General Virology

103

View full text Add to dashboard Cite

“…The gels were stained with 0.25% Coomassie blue R-250 in 50% ethanol, 15% acetic acid and destained in 10% ethanol, 10% acetic acid. Gels were also stained using Schiff's reagent prepared from basic fuchsin (Merck) according to the method of Segrest [29].…”

Section: Sds-pagementioning

confidence: 99%

Purification and characterization of a novel protein from bovine aorta that inhibits coagulation

Reutelingsperger¹,

Kop²,

Hornstra³

et al. 1988

European Journal of Biochemistry

View full text Add to dashboard Cite

A novel inhibitor of blood coagulation has been isolated from the intima of bovine aorta. The inhibitor, vascular anticoagulant (VAC), has been purified to an active fraction that contains two Coomassie-blue-staining bands (Mr = 34000 and M , = 32000, as judged by sodium dodecyl sulfate/polyacrylamide electrophoresis). Both bands are single-chain proteins, having no glycoprotein features. Furthermore, they do not contain any detectable 4-carboxyglutamic acid residues. Both proteins have an identical isoelectric pH of approximately 4.5. VAC binds in the presence of calcium ions to a bilayer consisting of 20% dioleoylglycerophosphoserine and 80% dioleoylglycerophosphocholine with a Kd = 6 nM. The binding is dependent on the calcium concentration: halfsaturation of binding occurs at a calcium concentration of 0.8 mM. The binding is completely reversible with EDTA. Furthermore the phospholipid/VAC ratio at saturation was n = 112 and n = 32 mol/mol for 0.5 mM Ca2+ and 2 mM Ca2+, respectively. Binding does not occur between VAC and pure dioleoylglycerophosphocholine. In a system with purified coagulation factors VAC inhibits the activation of prothrombin by factor X, and calcium only in the presence of negatively charged phospholipids. VAC decreases the V,,, and increases the K, of the factor-X,-catalyzed prothrombin activation. Based on these results, we conclude that we have purified from bovine aortic intima an anticoagulant protein, which exerts its activity through a calcium-dependent binding to negatively charged phospholipids, and thus interferes with the assembly of prothrombinase on the phospholipid surface.The enzymes active in the process of blood coagulation belong to the class of serine proteases, and circulate in the blood as nonenzymatic precursors, the zymogens [l -31. The activation of such a zymogen proceeds in a proteolytic process in which an already activated coagulation factor cleaves a specific bond of its substrate, a zymogen [4]. The rate of this reaction is extremely low, but can be greatly enhanced by the presence of certain nonenzymatic cofactors, which can be divided into protein cofactors and negatively charged surfaces. The factor-X,-catalyzed activation of prothrombin, for Correspondence to C. P. M. Reutelingsperger, Laboratorium voor Biochemie, Rijksuniversiteit Limburg, Postbus 616, NL-6200-MD Maastricht, The Netherlands Abbreviations. MPT test, modified prothombin time test; PhCOIle-Glu-Pip-Gly-Arg-NH-Np . HCI (S2337), N-benzoyl-~-isoleucyI-~-glutamyl-L-piperidyl-glycyl-L-arginine p-nitroanilide hydrochloride; DPhe-Pip-Arg-NH-Np . 2 HCI (S2238), ~-phenylakanyl-L-pipecolyI-L-arginine p-nitroanilide dihydrochloride; NaCl/Tris/HCl, 100 mM NaCI, 50 mM Tris/HCI, pH 7.5; NaCl/Tris/HCi/EDTA, NaCI/Tris/ HCI with 2 mM EDTA; NaCl/Tris/HCl/albumin, NaCl/Tris/HCl with 0.5 mg/ml human serum albumin, pH 7.9; RVV-X, purified factor X activator from Russell's viper venom; Gla, 4-carboxyglutamic acid; Ole2Gro-P-Cho, 1,2-dioleoyl-sn-glycero-3-phosphocholine; Ole2Gro-P-Ser, 1,2-dioleoyl-sn-glycero-3-ph...

show abstract

“…Sodium dodecy 1 sulphate (SDS)-polyacrylamide gel electrophoresis was carried out in glass tubes as described by Shapiro et al and pili samples were dissociated with SDS and dithiothreitol and prepared for electrophoresis as described by Wyckoff et al (1977). After electrophoresis, the gels were stained for protein with Coomassie brilliant blue R250 (Wyckoff et al, 1977), for glycoprotein with periodic acid-Schiff's reagent (Segrest & Jackson, 1972), for phosphoprotein using methyl green (Cutting & Roth, 1973) and for lipid with oil red 0 (Abodeely et aI., 1971). Densitometric scans of Coomassie blue-stained bands in the gels were carried out on a 'Quick Scan' densitometer (Helena Laboratories, Beaumont, Tex., U.S.A.) with a white light filter.…”

Section: E V E R Ymentioning

confidence: 99%

Purification of Pili from Bacteroides nodosus and an Examination of their Chemical, Physical and Serological Properties

Every

1979

Journal of General Microbiology

View full text Add to dashboard Cite

Pili from Bacteroides nodosus were purified to greater than 99 % homogeneity by precipitation at pH 4.0 and in MgCl, followed by chromatography on BioGel A150. The pili were composed entirely of one type of polypeptide subunit, pilin. No carbohydrates, nucleic acid, lipid, lipopolysaccharide or phosphate could be detected in purified pili preparations. The molecular weight of pilin from B. nodosus strains 9 1 B and 198 was 18 400 and from strain 80 was 19 300. The isoelectric points of pili from B. nodosus strains 91B and 80 were both 4.5.The buoyant densities of pili from strains 91B, 80 and 198 were 1.287, 1.284 and 1.286 g ml-l, respectively. The three strains of B. nodosus did not cross-react in K-agglutination tests and produced pili which did not cross-react in immunodiffusion tests. Antiserum to highly purified pili caused a characteristic K-type agglutination reaction. It was concluded that pili are the K-agglutinogen.

show abstract

[5] Molecular weight determination of glycoproteins by polyacrylamide gel electrophoresis in sodium dodecyl sulfate

Cited by 763 publications

References 7 publications

Oligomerization and post-translational processing of glycoprotein G of human respiratory syncytial virus: altered O-glycosylation in the presence of brefeldin A

Oligomerization and post-translational processing of glycoprotein G of human respiratory syncytial virus: altered O-glycosylation in the presence of brefeldin A

Purification and characterization of a novel protein from bovine aorta that inhibits coagulation

Purification of Pili from Bacteroides nodosus and an Examination of their Chemical, Physical and Serological Properties

Contact Info

Product

Resources

About