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J Kocourek

Publications and source records attributed to J Kocourek.

At least 19 recordsLinked to original sources
PubMed1979

Studies on lectins. XLIV. The pH dependence of lectin interactions with sugars as determined by affinity electrophoresis.

The pH dependence of association constants of the lectin-sugar complexes was determined by means of affinity electrophoresis. All the lectins studied (from the seeds of Dolichos biflorus, Glycine soja, Lens esculenta and Vicia cracca and of the fruiting body of Marasmius oreades) were characterized by a similar course of pH dependence of the association constants, with the maximum values at pH 7--9. For concanavalin A and the L-fucose binding Ulex europaeus lectin only the association constants at three selected pH values were determined. Concanavalin A does not interact with immobilized alpha-D-mannosyl residues at pH 2.3. The association constants vs. pH curves measured for lectins isolated from two different lentil varieties slightly differ in accordance with the differences observed in the interaction of these lectins with the Sephadex gel.

Binding Sites↗
PubMed1978

Studies on lectins. XXXIX. S-Glycosyl polyacrylamide gels for affinity chromatography of lectins.

Hydrophilic water-insoluble gels suitable for affinity chromatography of lectins have been prepared by copolymerization of acrylamide, N,N'-methylene bisacrylamide and alkenyl 1-thioglycosides. Water-soluble copolymers of analogous type have been obtained by omitting the cross-linking agent, N,N'-methylene bisacrylamide. In affinity chromatography of the Ricinus communis lectin it could be shown that the capacity for the lectin of the water insoluble copolymers was more than four times higher in copolymers having the S-beta-D-galactosyl ligand attached through a methylene bridge than in derivatives with a nonamethylene spacer. None of the insoluble S-beta-D-glycosyl copolymers prepared could be shown usable as affinity adsorbent for glycosidases though the corresponding soluble copolymers inhibited the activity of the enzymes.

Acrylamides↗
PubMed1978

Studies on lectins. XL. O-glycosyl derivatives of Spheron in affinity chromatography of lectins.

Free monosaccharides can be used for direct glycosylation of Spheron, a spherical macroporous hydroxyalkyl methacrylate-ethylene dimethacrylate copolymer, in a reaction that proceeds at room temperature in dioxane medium under catalysis of dry HCl or BF3. Derivatives of L-fucose, D-galactose, D-glucose, D-mannose, N-acetyl-D-galactosamine and N-acetyl-D-glucosamine thus prepared from Spheron beads have been shown to be efficient affinity carriers in isolation of lectins from seeds of Canavalia ensiformis D.C. (concanavalin A), Dolichos biflorus L., Glycine soja (L.) Sieb. et Zucc., Lens esculenta Moench, Ricinus communis L., Ulex europaeus L. and from albumin glands of the garden snail Helix pomatia L.

Acrylic Resins↗
PubMed1978

Studies on lectins. XLIII. Isolation and characterization of the lectin from restharrow roots (Ononis hircina Jacq.).

From 1 kg of dried Ononis hircina Jacq. roots 36 mg of a lectin were isolated by affinity chromatography on O-beta-lactosyl polyacrylamide gel. The lectin is homogeneous as judged by ultracentrifugal analysis (S20,W=6.2 S), polyacrylamide disc electrophoresis at pH 8.6 or 4.5, gel filtration on thin layers of Sephadex G-200 (Mr = 110 000) and dodecyl sulfate electrophoresis (Mr of subunits 31 000, both in presence and absence of mercaptoethanol) and disc dodecyl sulfate electrophoresis (pH 9.5). The lectin contains much aspartic and glutamic acids, serine and threonine and also 7.2% of neutral sugar. It is relatively specific for human type O erythrocytes that are agglutinated at a minimal lectin concentration 0.3 microgram/ml. The erythroagglutinating activity is not stimulated by Ca2+, Zn2+, Mg2+, Mn2+, Co2+ or Ni2+ salts; it is inhibited most effectively by N-acetyl-D-galactosamine and a number of D-galactose derivatives. Dissociation constants of several lectin . sugar complexes were estimated by affinity electrophoresis. The lectin is not mitogenic in rabbit lymph node lymphocytes.

ABO Blood-Group System↗
PubMed1978

The lack of insulin-binding to simple glycosidic receptors.

The results of affinity electrophoresis at pH 8.9 show that insulin does not interact with any of a number of glycosidic ligands which bind various lectins. The results are discussed with respect to some similarities between insulin and concanavalin A.

Binding Sites↗
PubMed1978

Studies on lectins. XLI. Isolation and characterization of a blood group B specific lectin from the role of the powan (Coregonus lavaretus maraena).

A B-specific lectin from the roe of the powan (Coregonus lavaretus maraena), a fish of the Salmonidae family, was isolated by affinity chromatography on O-alpha-D-galactosyl polyacrylamide gel. From 630 g of the lyophilized roe, 346 mg of pure lectin was obtained in a single isolation step. The lectin is electrophoretically homogeneous, its sedimentation coefficient s20,w is 2.9S and molecular weight 25 000. The molecular weight of the subunits estimated by electrophoresis in the presence of dodecyl sulfate is 27 000 for both reduced and nonreduced substance. The lectin contains a large amount of cysteine, has a small content of aromatic amino acids, 10.4% of neutral sugar and 0.145% of Zn. It agglutinates specifically human B-group erythrocytes; the agglutination is stimulated by Zn2+ and Mg2+ ions and partially inhibited by EDTA. The most efficient carbohydrate inhibitors are methyl alpha-L-rhamnoside (6 micrometer), L-rhamnose (12 micrometer) and raffinose (0.1 mM). The association constant of the complex lectin . L-rhamnose is KA = 9.5 . 10(2) M-1, as determined by fluorimetric titration.

ABO Blood-Group System↗
PubMed1978

Studies on lectins. XXXVII. Isolation and characterization of the lectin from Jimson-weed seeds (Datura stramonium L.).

The lectin of Jimson-weed seeds (Datura stramonium L.) was isolated by affinity chromatography on a polysaccharide mixture from mycelium of Aspergillus niger. The lectin yields two bands on disc electrophoresis, it has sedimentation coefficient s20,w = 3.8 S and its apparent molecular weight estimated by thin layer gel chromatography is 120,000. The lectin reduced with mercaptoethanol yields on polyacrylamide gel electrophoresis in the presence of dodecyl sulfate three zones corresponding to subunits of molecular weight 72,000, 45,000 and 25,000. The lectin contains large amounts of cystine, glycine, 6.3% of hydroxyproline residues, 4.5% glucosamine and 28% of neutral sugar, predominantly arabinose. The lectin is nonspecific in human erythrocyte ABO system, it is not inhibited by simple sugars but is inhibited by a partial hydrolysate of chitin-containing mixture of polysaccharides from Aspergillus niger.

Amino Acids↗
PubMed1978

Studies on lectins. XXXVIII. Isolation and characterization of the lectin from black locust bark (Robinia pseudacacia L.).

The lectin of black locust (Robinia pseudacacia) bark was isolated by specific adsorption on formaldehyde-fixed human erythrocytes and elution with a borate solution. The lectin is homogeneous on disc electrophoresis and ultracentrifugation (s20,w = 5.8 S) but yields three bands on isoelectric focusing. It has a molecular weight of approximately 110,000 and consists of two types of subunit (mol. wt 29,000 and 31,500). Its pI is approximately 5.9; it contains high amounts of aspartic acid, threonine and serine, no cysteine and very little methionine. Also 7.2% of covalently bound neutral sugar and 0.47% of glucosamine are present. The lectin is nonspecific in agglutination of human erythrocytes, it is inhibited by high concentrations of N-acetyl-D-galactosamine and is mitogenic in rabbit lymph node lymphocytes.

Amino Acids↗
PubMed1978

Studies on lectins. XXXV. Water-soluble O-glycosyl polyacrylamide derivatives for specific precipitation of lectins.

By copolymerization of acrylamide and allyl glycosides of various sugars, O-glycosyl derivatives of polyacrylamide copolymers were prepared. The sugar content of the copolymers can be varied in the range 0--40%, their sedimentation coefficient shows the vales of 2.5-5.7 S; the molecular weight of an O-alpha-D-mannopyranosyl polyacrylamide copolymer (29% mannose, so20,w = 2.9 S) was estimated as 44 500. Copolymers with incorporated glycosyl residues interacting specifically with lectins yield precipitates with them upon immunodiffusion in cellulose acetate. The quantitative precipitin curves obtained with these copolymers are similar to those produced by quantitative precipitation of lectins with natural polysaccharides. The copolymers may serve as model substances of natural polysaccharides.

Acrylamides↗
PubMed1978

Studies on lectins. XXXVI. Properties of some lectins prepared by affinity chromatography on O-glycosyl polyacrylamide gels.

A number of lectins has been purified by affinity chromatography on O-glycosyl polyacrylamide gels. The lectins isolated (and the particular sugar ligands used in the affinity carriers) are as follows: Anguilla anguilla, serum (alpha-L-fucosyl-), Vicia cracca, seeds; Phaseolus lunatus, seeds; Glycine soja, seeds; Dolichos biflorus, seeds; Maclura pomifera, seeds; Sarothamnus scoparius, seeds; Helix pomatia, ablumin glands; Clitocybe nebularis, fruiting bodies (all N-acetyl-alpha-D-galactosaminyl-); Ricinus communis, seeds (beta-lactosyl-); Ononis spinosa, root; Fomes fomentarius, fruiting bodies; Marasmius oreades, fruiting bodies (all alpha-D-galactosyl-), Canavalia ensiformis, seeds, (i.e., concanavalin A) (alpha-D-glucosyl-). Physicochemical properties of Glycine soja, Dolichos biflorus, Phaseolus lunatus, Helix Pomatia and Ricinus communis lectins corresponded well to properties of the preparations studied earlier by other workers. For the other purified lectins the essential physiochemical data (sedimentation coefficient, molecular weight, subunit composition, electrophoretic patterns, amino acid composition, carbohydrate content, isoelectric point) were established and their precipitating, hemagglutinating and mitogenic activities determined.

Agglutinins↗
PubMed1978

Additional precipitation reactions of lectins with human serum glycoproteins.

Highly purified human serum glycoproteins were treated with neuraminidase and examined for their cross reaction with several lectins with anti-galactosyl specificity: beta-D-galactosyl structures are thought to be the main terminal sugar residues that become attached de novo after removal of neuraminic acid. The following lectins were tested: Tridacnin from the bivalve clams Tridacna maxima and Tridacna gigas, the agglutinin from the sponge Axinella polypoides, the lectin from the roach Rutilus rutilus and the plant lectins from Ricinus communis, Ononis spinosa, Glycine soja and Abrus precatorius. In agar gel diffusion, these purified and precipitating lectins gave more or less strong or negative results against the different neuraminidase-treated serum glycoproteins, thus indicating subtle differences with respect to their anti-galactosyl combining specificity. On the other hand, serum glycoproteins which reacted with the same lectin, did not always show complete identity lines. Finally, as revealed by these lectins, the carbohydrate moiety of serum glycoproteins may reflect a complex and broad spectrum of heterogeneity. This could lead to a more detailed understanding of the topographical and steric arrangement of the chemical structure and of the biological role of carbohydrate groups in these glycosubstances.

Animals↗
PubMed1978

Studies on lectins. XXXIV. The effect of thermal treatment on the lectin of lentil seeds (Lens esuclenta Moench).

The lectin of lentil seeds (Lens esculenta Moench) dissociates into subunits upon incubation of its solution at temperatures above 50 degrees C. The dissociation is accompanied by precipitation of the heavy subunit, release of Mn2+ from the protein molecule and an almost complete loss of erythroagglutinating activity. Addition of Mn2+ or D-glucose to the lectin solutions has a partially protective effect on the dissociation and inactivation.

Glucose↗
PubMed1977

Studies on lectins. XXXI. Determination of dissociation constants of lectin. Sugar complexes by means of affinity electrophoresis.

A modification of affinity electrophoresis in polyacrylamide gels containing immobilized sugar residues is described. The immobilization of sugar residues is achieved by addition of a water soluble O-glycosyl polyacrylamide copolymer to the polymerization mixture, which serves for the preparation of gels for commonly used discontinuous polyacrylamide gel electrophoresis. The electrophoretic mobility of lectins in affinity gels containing specific immobilized sugar decreases with increasing sugar concentration. The addition of free sugar into the affinity gel abolishes retardation of the electrophoretic mobility caused by immobilized sugar. The relationship between electrophoretic mobility of lectins and concentration of immobilized and free sugar was used for the calculation of dissociation constants of the various complexes lectin-immobilized sugar and lectin-free sugar.

Electrophoresis, Polyacrylamide Gel↗
PubMed1977

Studies on lectins. XXXII. Application of affinity electrophoresis to the study of the interaction of lectins and their derivatives with sugars.

Affinity electrophoresis was used to study the sugar binding heterogeneity of lectins or their derivatives. Commercial and demetallized preparations of concanavalin A could be resolved by affinity electrophoresis into three components with different affinity to immobilized sugar. Similarly the Vicia cracca lectin obtained by affinity chromatography behaved on affinity gels as a mixture of active and inactive molecular species. Affinity electrophoresis has shown that the nonhemagglutinating acetylated lentil lectin and photo-oxidized or sulfenylated pea lectin retain their sugar binding properties; dissociation constants of saccharide complexes of these derivatives are similar to those of native lectins. The presence of specific immobilized sugar in the affinity gel improved the resolution of isolectins from Dolichos biflorus and Ricinus communis seeds.

Acetylation↗
PubMed1977

alpha-Mannosidase and mannanase of some wood-rotting fungi.

Cultivation media from 11 wood-rotting fungi contained alpha-mannosidase and mannanase activity, alpha-Mannosidase was studied in more detail in Phellinus abietis and mannanase was studied more intimately in basidiomycetes Phellinus abietis, Trametes sanguinea and Pholiota aurivella. Suitable cultivation conditions and optimum conditions for the production of alpha-mannosidase and mannanase were determined. Both enzymes are constitutive; mannanase is extracellular, alpha-mannosidase was found in both mycelium and cultivation medium.

Basidiomycota↗
PubMed1977

Separation and properties of alpha-mannosidase and mannanase from the basidiomycete Phellinus abietis.

Proteins of a crude enzyme preparation obtained from the cultivation medium of the basidiomycete Phellinus abietis were separated by gel filtration and ion-exchange chromatography. The preparation contained a minimum of three enzymes capable of splitting alpha-D-mannosidic bonds: alpha-mannosidase, exomannanase, and endomannanase, which were separated. Some properties of the mannanase complex of the crude enzyme preparation, and of a partially purified alpha-mannosidase were examined. The mannanase complex exhibited two pH optima, its temperature optimum being at 45 degrees C. The pH optimum of purified alpha-mannosidase was at pH 5.0, the temperature optimum being at 45 degrees C. The pH optimum of purifed alpha-mannosidase was at pH 5.0, the temperature optimum at at 60 degrees C; the enzyme had a relatively high heat stability. The Km of alpha-mannosidase for p-nitrophenyl alpha-D-mannopyranoside was 1.5 X 10(-5) M. Pure alpha-mannosidase did not split mannan.

Basidiomycota↗