Montri Chulavatnatol

Characterization of cyanogenic β-glucosidase (Linamarase) from cassava (Manihot esculenta Crantz)☆

Abstract Linamarase (EC 3.2.1.21) was purified from cassava petiole, stem, and root cortex by ammonium sulfate precipitation, column chromatography on Sepharose 6B, and chromatofocusing. The last step resolved the enzyme from each source into three forms with pI values of 4.3, 3.3, and 2.9. Each form was found to be oligomeric, consisting of one kind of subunit, Mr 63,000. The major isozyme with a pI of 4.3 from petiole showed a Km for linamarin of 0.6 m m and possessed both β-glucosidase and β-fucosidase activities. The former was sensitive to inhibition by δ-gluconolactone, isopropyl-β- d -thioglucoside, and HgCl2, whereas the latter was inhibited by Tris ion.

Interaction of a mulberry leaf lectin with a phytopathogenic bacterium, P. syringae pv mori.

Two N-glycolylneuraminic acid-specific lectins, MLL 1 and 2, from leaves of Morus alba were studied for their anti-bacterial activity against P. syringae pv mori, which was a specific pathogenic bacterium of the mulberry leaf. MLL 1 but not MLL 2 was found to induce the agglutination of P. syringae pv mori. The MLL 1 can induce the agglutination only at the exponential phase of the bacterial growth in a liquid medium and the agglutination was specifically inhibited by N-glycolylneuraminic acid, N-acetylgalactosamine at 12.5 mM and bovine submaxillary mucin at 0.05 &mgr;g/ml.

Monodin-induced agglutination of Vibrio vulnificus, a major infective bacterium in black tiger prawn (Penaeus monodon)

Abstract 1. 1. Monodin, a sialic acid-specific lectin from P. monodon, was found to induce the agglutination of Vibrio vulnificus, a major infective bacterium of the prawn. 2. 2. The monodin-induced agglutination of V.vulnificus was specifically inhibited by N-acetylneuraminic acid and anti-monodin. 3. 3. During growth in a liquid medium, V. vulnificus can be induced to agglutinate only during the early stationary phase. 4. 4. An elevation of the monodin level was found in most of the prawns suffering from bacterial infection.

Monodin, a new sialic acid-specific lectin from black tiger prawn (Peneaus monodon)

1. 1. A new lectin, called monodin, was purified from the hemolymph of Peneaus monodon by affinity chromatography using a fetuin-agarose column. 2. 2. Monodin is a glycoprotein, having a native mol. wt of 420,000 and a subunit mol. wt of 27,000. 3. 3. The monodin activity was specific for N-acetylneuraminic acid and required 1 mM calcium ion. 4. 4. The antibody to monodin interacted with the extracts of various prawn tissues and serum of other prawns.

Sialic acid binding lectins from leaf of mulberry (Morus alba)

Abstract Two new lectins, MLL 1 and MLL 2, were purified from the leaves of mulberry ( Morus alba ) by a procedure consisting of ammonium sulfate precipitation, affinity chromatography on a N -acetylgalactosamine-agarose column, gel filtration on a Sephacryl S-200 column, anion-exchange chromatography using a DEAE-Sephacel column and gel filtration on a Sephadex G-75 column. Both MLL 1 and MLL 2 were found to be glycoproteins with neutral sugar contents of 8.8 and 40%, respectively. They showed the same native molecular weight of 51 000. Each consisted of subunit with molecular weight of 16 500. They were highly specific for N -glycolylneuraminic acid.

Effects of ATP, cAMP and pH on the initiation of flagellar movement in demembranated models of rat epididymal spermatozoa☆

Abstract Demembranated model of rat epididymal spermatozoa was employed to establish the conditions for the initiation of flagellar movement. Extensive initiation of the flagellar movement required 0.5 mM ATP, 1 μM cAMP and pH 7.9. The requirement for ATP was highly specific and can partially be replaced by 2′-deoxy-ATP only, but not by analogs of ATP or other nucleoside triphosphates. In contrast, the cAMP requirement was less specific and can partially be replaced by other cyclic nucleotides and cAMP-analogs except 2′-deoxy-cAMP and 2′,3′-cAMP. The data implied that the intracellular pH rise, not the cAMP increase, was the probable trigger for the initiation of sperm motility after ejaculation. During sperm maturation, the sperm motile apparatus appeared unchanged with respect to the above conditions of reactivation.

A sperm-agglutinating lectin from seeds of jack fruit (Artocarpusheterophyllus)

A lectin specific for N-acetylgalactosamine was isolated from seed extract of Jack fruit (Artocarpus heterophyllus) by ammonium sulfate precipitation, followed by affinity chromatography on a Affigel-galactosamine-agarose column. The lectin possessed agglutinating activities for human and rat sperm as well as human red blood cells. It was found to have Mr = 62,000 consisting of two dissimilar subunits of Mr = 18,000 and 13,000. It also cross-reacted with an antibody against the lectin of Osage Orange (Maclura pomifera).

A new acidic protease in human seminal plasma

Summary A new acidic protease with pH optimum of 3.0 was partially purified from human seminal plasma. It can hydrolyze hemoglobin and N, N-dimethyl casein. It was stable in acidic but labile in neutral and alkaline solutions. Its molecular weight was determined by gel filtration to be 40,000. Its chromatographic behavior on carboxy methyl Sephadex column was quite different from the pH-8.0 protease of human seminal plasma. It was present in seminal plasma of normal and vasectomized individuals. Its activity was not affected by cysteine, metal cations or ethylene diamine tetraacetic acid.

Activation of proenzyme of acidic protease from human seminal plasma

The kinetics and the extent of the conversion of the proenzyme into the active acidic protease (EC 3.4.23.--) of human seminal plasma were dependent on acidic pH. Between pH 2 and 4, the initial rate of the activation was first-order with respect to the proenzyme. Between pH 4.5 and 5, the rate deviated from the first-order with an initial lag period which can be abolished by adding an excess amount of the acidic protease or pepsin. The extent of the activation was complete between pH 2 and 3 and became incomplete between pH 4 and 5. Addition of the acidic protease or pepsin did not alter the extent of the activation at the high pH values. According to the chromatographic profile on a Sephadex G-75 column, the activation products (namely active acidic protease and an activation peptide) obtained at pH 3 and those obtained at pH 4.5 were identical. The molecular weight of the activation peptide obtained at pH 3 was 6900; its amino acid composition was analyzed and compared with those of the proenzyme and the acidic protease. Remarkable similarity between the amino acid composition of the acidic protease and that of human pepsin was observed. In the presence of an excess amount of hemoglobin, the conversion of the proenzyme was self-activated and showed an initial lag period. Addition of acidic protease did not change the rate of self activation or the lag period.

Acid protease and its Proenzyme from Human Seminal Plasma

An acid protease, with an optimum pH of 2.5, exists in seminal plasma in a proenzyme form. In acidic pH, the proenzyme is converted into the active form, resulting in the release of small molecular weight peptide. The extent and rate of proenzyme-active enzyme conversion is absolutely dependent on pH. Between pH 5 and 2, as the pH is lowered, the extent of conversion increases and reaches a maximum between pH 3 and 2. The kinetics of activation shift from first-order between pH 2 and 4 to more complexity with a lag period between pH 4.5 and 5. Under physiological conditions, the proenzyme might be activated by coming into contact with acidic vaginal fluid during ejaculation. The acid protease can hydrolyze the cervical mucus protein.