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J Kocourek

Publications and source records attributed to J Kocourek.

24 records · Page 2Linked to original sources
PubMed1976

Studies on phytohemagglutinins. XXVIII. Chemical modifications of the lectin from seeds of the lentil (Lens esculenta Moench.)

The reaction of lentil lectin with maleic anhydride yields maleyl derivatives with a degree of modification depending on the amount of used maleic anhydride. Hemagglutinating activity of maleyl derivatives decreases with the increasing number of modified lysine residues, but even at a maleylation of 90% of free -NH2 groups, the maleyl derivative of the lectin still retains a hemagglutinating activity. Maleylation of the lentil lectin is not accompanied by dissociation; the chemically modified lectin also retains its bound Mn. Maleyl derivatives of the lectin retain the ability to interact specifically with polysaccharides (Sephadex, mannan). After removal of maleyl groups in acidic medium, the hemagglutinating activity does not differ from that of the native lectin. Acetylation of the lentil lectin with N-acetylimidazole results in formation of a modified protein containing 3-5 acetylated -OH groups of the tyrosyl residues out of the total of 15 groups and 7-17 acetylated -NH2 groups out of the total of 26 free -NH2 groups, depending upon the reaction conditions. The presence of D-mannose in the reaction mixture does not affect the extent of modification of tyrosyl groups, but it decreases the modification of -NH2 groups. Acetyl derivatives of the lentil lectin do not possess hemagglutinating activity, but they retain the ability to interact with polysaccharides and to bind to erythrocytes. Acetylation does not affect the content of protein bound Mn. After de-O-acetylation, the hemagglutinating activity of the modified protein increases to the value of the native protein.

Acetates↗
PubMed1976

Studies on phytohemagglutinins. XXVII. A study of the pea lectin binding site.

Under defined mild conditions the reaction of the pea lectin with 2-nitrophenylsulfenyl chloride results in sulfenylation of only 2 of the 10 tryptophan residues of the lectin molecule with simultaneous loss of biological activity. Both sulfenylated tryptophan residues belong to the two heavy subunits of the lectin. Enzymic hydrolysis and separation of the tryptic peptides yields only one homogeneous yellow peptide containing the modified tryptophan residue. The isolated peptide has the following sequence (NPS, nitrophenylsulfenyl): HAsp-Val-Val-Pro-Glu-(2-NPS-Trp)-Val-ArgOH. The octapeptide is either directly a part of the pea lectin binding site or it plays an important role in maintaining the tertiary structure of the binding site. According to the amino acid composition and amino acid sequence, the octapeptide isolated from the pea lectin is almost identical with that part of the peptide chain of concanavalin A near to which the location of the sugar binding site is supposed to be.

Amino Acid Sequence↗
PubMed1975

Studies on phytohemagglutinins. XXV. Isolation and characterization of hemagglutinins of the spindle tree seeds (Evonymus europaea L.).

A mixture of isophytohemagglutinins has been isolated from the fleshy arils of the spindle tree seeds (Evonymus europaea L.) by fractional precipitation of the saline extract of the arils by (NH4)2SO4 at a 0.40% saturation. Successive preparative disc electrophoresis on polyacrylamide gel affords separation of one slower moving component, phytohemagglutinin I, from the mixture of other isophytohemagglutinins that have a very similar electrophoretic mobility. Phytohemagglutinin I has a sedimentation coefficient Sw,20 of 7.1 S and an approximate mol. wt of 127 000. Amino acid analysis shows a high amount of aspartic acid, alanine and glycine but also significant amounts of serine, threonine, cysteine and methionine. Aspartic acid is the only N-terminal amino acid found by the dansylation technique. Phytohemagglutinin I contains glucosamine and 4.7% neutral sugar. Its approximate pI in citrate/phosphate buffer is 4.4-4.5. The metal content amounts to 0.250% Ca, 0.019% Mg, 0.034% Zn and 0.026% Cu. Mn is not present. Ultracentrifugation analysis reveals homogeneity in the sedimentation behavior of the mixture of isophytohemagglutinin, an Sw,20 of 7.1 S and an approximate mol. wt of 119 000. The mixture has an amino acid composition closely resembling that of phytohemagglutinin I and an identical pI but contains only 1.9% neutral sugar. Two N-terminal amino acids were shown to be present, aspartic acid and tyrosine. With the exception of Cu which is absent, the metal content is almost the same as that of phytohemagglutinin I. Both phytohemagglutinin I and the mixture are devoid of anti-A1 activity and show detectable anti-H, anti-B and anti-A2 erythroagglutinating activity in approximate limit concentrations of 2.5, 5 and 10 mug/ml, respectively. This activity is not influenced by the presence of EDTA, Ca2+ or Mg2+, but is stimulated by Zn2+. Mn2+ and Co2+ have an inhibitory effect. None of the simple sugars tested inhibited the hemagglutination reactions.

Amino Acids↗
PubMed1975

Studies on phytohemagglutinins. XXVI. A new type of a lentil hemagglutinin isolated from Lens esculenta Moench., subsp. microsperma (Baumg.) Barulina.

In seeds of the lentil Lens esculenta Moench., subsp. microsperma (Baumg.) Barulina, a nonspecific phytohemagglutinin was found, the properties of which differ from those of the previously described lentil hemagglutinins. The phytohemagglutinin is not adsorbed to the Sephadex matrix; its hemagglutinating activity is different towards erythrocytes of different human blood groups of the ABO system. The isolated phytohemagglutinin preparation is homogeneous by disc polyacrylamide and starch gel electrophoreses and by gel chromatography on Bio-Gel P-100. Ultracentrifugation of the phytohemagglutinin yields a single symmetrical peak with S20,W of 3.9 S. From the sedimentation data, a molecular weight of 53 300 was calculated. Phytohemagglutinins of similar properties were shown to be present also in other cultivars of Lens esculenta, subsp. microsperma.

ABO Blood-Group System↗
PubMed1975

Studies on phytohemagglutinins. XXIV. Isoelectric point and hybridization of the pea (Pisum sativum L.) isophytohemagglutinins.

The isoelectric point of the two pea isophytohemagglutinins varies from pH 5.7 to pH 8.4 depending on the composition of the buffer used. Isoelectric focusing reveals three main molecular species with pI at pH 5.90, 6.35 and 7.00. Molecular species with pI at pH 5.9 and 7.0 correspond to the two pea isophytohemagglutinins which can be obtained by DEAE-cellulose chromatography (Entlicher, G., Kostír, J.V. and Kocurek, J. (1970) Biochim. Biophys. Acta 221, 272-281). A molecular species with pI at pH 6.35 is formed from the two pea isophytohemagglutinins by hybridization. According to the hybridization pattern and subunit composition of the pea isophytohemagglutinins the subunit composition AABB, AACC and AABC can be proposed for the three molecular species with respect to ionic properties of the subunits.

Buffers↗