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J Kocourek

Publications and source records attributed to J Kocourek.

At least 37 records · Page 2Linked to original sources
PubMed1983

Studies on lectins. LVII. Immunofluorescence localization of lectins present in fish ovaries.

The occurrence of endogeneous lectins in the ovaries of four fish species has been studied by indirect immunofluorescence staining with antibodies against individual lectins. Paraffin sections of the ovary of perch (Perca fluviatilis L.) were treated with an antibody against perch lectin. In cryostat sections of the tench (Tinca tinca L.) ovary, the L-rhamnose-specific lectin "I" was detected with a specific antibody. In cryostat sections of both roach (Rutilus rutilus L.) and rudd (Scardinius erythrophthalmus L.) ovaries, lectins were localized using a single antibody against roach lectin. The isolation of tench lectins is briefly described. In the fish species employed for this study, lectins are associated exclusively with the content and surrounding membrane of cortical vesicles situated within the cytoplasm of maturing oocytes. The positive reaction with lectin antibody was observed almost immediately after the formation of the first cortical vesicles in the peripheral cytoplasm of early previtellogenic oocytes. Their lectin content increases during the later stages when cortical granules fill the whole cytoplasm before moving towards the cell periphery, as the oocyte starts to accumulate yolk. The presence of lectins within cortical vesicles is significant also in view of the polysaccharide content of these structures. In the vitellogenic oocytes lectins seem to move towards the cell periphery and accumulate beneath the plasma membrane. Our observations are discussed in view of the present ideas on the intracellular function of lectins, and with respect to the role of cortical vesicles in fertilization and in post-fertilization modifications of the egg envelopes.

Animals↗
PubMed1982

Synthetic glycosyl polymers in isolation, characterization and immobilization of lectins.

A review is given on preparative procedures and practical applications in the field of lectins of two types of synthetic polymers: (1) Glycosyl polyacrylamide derivatives - insoluble (cross-linked) and water-soluble (linear) - that are obtained by copolymerization of alkenyl glycosides and acrylamide, and (2) glycosyloxyethyl dimethacrylate copolymers prepared by glycosylation of the commercially available macroporous glycol methacrylate copolymer, Separon. Both types of copolymers are efficient affinity carriers for isolation of lectins. Separon derivatives can be used after partial periodate oxidation of the glycosyl groups for covalent coupling of lectins. Affinity carriers thus obtained can be employed in separation and purification of glycoproteins and other glycosylated macromolecules. Soluble glycosyl polyacrylamide derivatives are useful in precipitation of lectins and, in general, as synthetic model substances for studying interactions of lectins with sugar ligands.

Acrylic Resins↗
PubMed1982

Studies on lectins. LII. Isolation and characterization of the lectin from rye germ (Secale cereale L.).

Rye germ lectin was isolated by extraction of defatted rye germ, fractionation of the extract by ammonium sulfate precipitation, affinity chromatography of the active substances on chitin-beta-glucan and gel filtration on Sephadex G-50. The lectin shows erythroagglutinating activity at a minimum concentration of 2.5 micrograms/ml. The erythroagglutinating activity is the same against human red blood cells of all types of the ABO system and is inhibited by N-acetyl-D-glucosamine. The lectin has no mitogenic activity against mouse splenic lymphocytes. According to the results of polyacrylamide gel electrophoresis the lectin obtained is a mixture of three very similar isolectins of equal erythroagglutinating activity. Sedimentation analysis indicates homogeneity of the lectin preparation; the molecular weight was 56000 as estimated by sedimentation equilibrium. In the presence of urea and sodium dodecyl sulfate the lectin dissociates into 2 types of subunits with molecular weights of 35000 and 19000. The rye germ lectin contains about 2% of neutral sugar and 1% of D-glucosamine. The amino acid composition of the lectin is characterized by a very high content of glycine and half cystine and a low content of apolar amino acids. N-terminal amino acids of the lectin are apparently blocked.

Acetylglucosamine↗
PubMed1981

Studies on lectins. LI. The role of Mn2+ in the activity of the soybean lectin.

In contradistinction to previous reports, the lectin of soybeans (Glycine soja) has been shown to retain its erythroagglutinating activity after complete removal of manganese from its molecule. The applied demetallization procedures (dialysis against 0.1 M HCl or 1M acetic acid and against 0.1 M EDTA following by dialysis against 1 M acetic acid) had no effect on the stability of the lectin subunit structure, as proved by ultracentrifugation analysis or thin-layer chromatography on Sephadex G-200 Superfine. Affinity electrophoresis on polyacrylamide gel containing immobilized alpha-D-galactosyl residues and affinity chromatography on alpha-D-galactosyl derivative of Separon have shown that the interaction of the demetallized lectin preparations does not differ from that of the native lectin in alkaline media but is decreased in acidic media. Demetallized preparations of soybean lectin can be fully reactivated by dialysis against solutions containing Mn2+, Zn2+ or Cd2+.

Animals↗
PubMed1981

Isolation and properties of a lectin from the seeds of hairy vetch (Vicia villosa Roth).

The lectin of the seeds of hairy vetch (Vicia villosa Roth), which selectively binds murine cytotoxic T-lymphocytes, was purified by simple affinity-chromatographic procedures on two different N-acetyl-alpha-D-galactosaminyl-carriers. The lectin thus obtained is homogeneous on polyacrylamide-gel electrophoresis both in acid and alkaline media and has a mol. wt. of approx. 120000. The lectin molecule appears to comprise four subunits of equal electrophoretic mobility, contains 4.3% of covalently bound neutral sugar and 0.72 Mn and 0.94 Zn atoms respectively. The anti-(blood-group A1) specific erythroagglutinating activity of the lectin can be detected at a limit concentration of 15 microgram/ml and is inhibitable most effectively by N-acetyl-D-galactosamine.

Amino Acids↗
PubMed1980

Studies on lectins. XLVII. Some properties of D-galactose binding lectins isolated from the seeds of Butea frondosa, Erythrina indica and Momordica charantia.

The lectins from the seeds of Butea frondosa, Erythrina indica and Momordica charantia were isolated by affinity chromatography on O-alpha-D-galactosyl polyacrylamide gels. The Butea frondosa lectin has sedimentation coefficient S20,W = 6.7 S, its molecular weight is 141 000 and it consists of noncovalently associated subunits of molecular weights 36 000 and 65 000. The lectin contains two 1/2-cystine and 3--4 methionine residues per molecule, 4.3% neutral sugar and 0.5% glucosamine. Threonine is the single N-terminal amino acid residue. The lectin from Erythrina indica seeds has a sedimentation coefficient S20,W = 4.0 S, its molecular weight is 66 200 and it consists of two kinds of subunit (molecular weights 30 000 and 34 000, respectively), which are noncovalently associated. The lectin is devoid of cystein, has a neutral sugar content 5.6% and glucosamine content 0.46%. It contains probably one Mn atom per subunit. The N-terminal amino acid sequence is Val-Glu-Val-Leu-(Phe)-Phe-(Ala)-Phe-. The lectin from Momordica charantia seeds has a sedimentation coefficient S20,W = 6.5 S, its molecular weight is 129 000. It interacts weakly with Sephadex G-200. The lectin consists of subunits of molecular weights 29 000, 32 000 and 36 000, respectively, held together by cystine bridges. Its content of cystine and methionine residues is relatively high, 4.1% neutral sugar and 0.5% glucosamine form the carbohydrate moiety. Two N-terminal amino acids (asparagine and valine) are present.

Amino Acids↗
PubMed1980

Studies on lectins. XLIX. The use of glycosyl derivatives of Dextran T-500 for affinity electrophoresis of lectins.

p-Aminophenyl glycosides and glycosylamines were coupled to periodate oxidized Dextran T-500 either directly or through an epsilon-aminocaproic acid spacer. The new glycosylated derivatives of dextran specifically precipitate lectins having the appropriate carbohydrate specificity, and thus were used in the preparation of affinity gels for affinity electrophoresis of lectins. The apparent strength of interaction of several lectins with carbohydrate residues immobilized in this way was less than with carbohydrates immobilized in O-glycosyl polyacrylamide copolymers. The presence of epsilon-aminocaproic spacer had no effect on the strength of interaction. The advantages of this type of macromolecular derivative of the ligand for affinity electrophoresis and some differences between the glycosylated dextrans and O-glycosyl polyacrylamide copolymers are discussed. Dextrans containing bound p-aminophenyl alpha-D-mannopyranoside and p-aminophenyl alpha-D-glucopyranoside were used to study the binding properties of concanavalin A and the lectin from Lathyrus sativus seeds. For the investigation of interaction of lectins from Ricinus communis and Glycine soja seeds, dextran derivatives containing bound p-aminophenyl alpha- and beta-D-galactopyranosides and alpha- and beta-D-galactopyranosylamines were used.

Binding Sites↗
PubMed1980

Studies on lectins. XLVIII. Isolation and characterization of lectins from the seeds of Lathyrus odoratus L. and Lathyrus silvestris L.

Lectins from seeds of Lathyrus odoratus L. and Lathyrus silvestris L. were isolated by (NH4)2SO4 precipitation and affinity chromatography on Sephadex G-150. Both lectins were found to be homogenous by ultracentrifugation (S20,w for the L. odoratus lectin was 3.29, for the L. silvestris lectin 3.39) and by discontinuous polyacrylamide gel electrophoresis in alkaline and acidic buffer systems. The amino acid composition of both lectins is characterized by high contents of aspartic and glutamic acids and threonine. The lectin from L. odoratus seeds contains 2.5% of neutral sugar, and the lectin from L. silvestris seeds 3.1%. Amino sugars were not detected. Threonine and valine were determined as N-terminal amino acids in the case of both lectins. Both lectins contain Mn in an amount corresponding approximately to 2 atoms per molecule. The molecular weight of the isolated lectins is 53 000, as determined by thin layer gel chromatography. Both lectins consist of two types of subunits differing in their molecular weights. Both Lathyrus lectins are nonspecific in agglutination of human erythrocytes; their hemagglutinating activity is inhibited by D-mannose and D-glucose and their alpha-glycosides. The dissociation contants of complexes of the studied lectins with D-glucose and D-mannose were determined using affinity electrophoresis.

Amino Acids↗
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↗