Mannose-specific lectin isolation from Canavalia ensiformis seeds by PHEMA-based cryogel.

Mannose-specific lectin Concanavalin A (Con A) was purified from Canavalia ensiformis seeds. For this purpose, mannose attached poly(hydroxyethyl methacrylate) (PHEMA) cryogel was prepared by cryopolymerization. Mannose was used as the affinity ligand and was covalently attached onto the PHEMA cryogel via carbodiimide activation. The PHEMA cryogel containing 23.3 mmol mannose/g polymer were used in the binding studies. Con A binding with the mannose attached PHEMA cryogel from Con A aqueous solution was 5.2 mg/g at pH 7. Maximum binding capacity for Con A from C. ensiformis seed extract was 39 mg/g. Con A was eluted with 0.3 M galactose, and the purity of Con A was determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. It was observed that the mannose attached PHEMA cryogel can be used without significant decrease in Con A binding capacity after six binding-elution cycles.

ConBr, a lectin from Canavalia brasiliensis seeds, protects against quinolinic acid-induced seizures in mice.

Lectins are proteins capable of reversible binding to carbohydrates or glycoconjugates. In the central nervous system of mammals, lectins with affinity for mannose/glucose or galactose can modulate cellular communication. ConBr, a lectin isolated from the seeds of Canavalia brasiliensis, previously showed antidepressant effect in the forced swimming test in mice, with involvement of the monoaminergic system. In this study, we investigated the neuroprotective effects of ConBr against quinolinic acid (QA), a well-known NMDA agonist that produces severe neurotoxicity when administered in vivo. ConBr (10 µg/site) administered via intracerebroventricular (i.c.v.) showed a neuroprotective activity against seizures induced by QA (36.8 nmol/site; i.c.v.) when administered 15 min prior to QA, with a percentage of protection around 50%. ConBr was also able to significantly decrease the severity of the seizures but without changes in the latency of the first convulsion or the duration of the seizures. This effect was dependent on the structural integrity of the ConBr protein and its binding capacity to oligosaccharides residues. ConA, a lectin with high similarity to ConBr, did not reverse the QA-induced seizures. Moreover, ConBr was able to protect against hippocampal cell death caused by QA, which was measured by propidium iodide incorporation. QA caused activation of JNK2 and improved the phosphorylation of Ser831 and 845 on the AMPA receptor GluR1 subunit, and both of these effects were counteracted by ConBr. Our data suggest that the lectin ConBr may exert a modulatory action on NMDA receptors, which inhibits its activity in response to QA.

Inhibition of jack bean urease by tetrachloro-o-benzoquinone and tetrachloro-p-benzoquinone.

Tetrachloro-o-benzoquinone (TCoBQ) and tetrachloro-p-benzoquinone (TCpBQ) were studied as inhibitors of jack bean urease in 20 mM phosphate buffer, pH 7.0, 1 mM EDTA, 25 degrees C. The mechanisms of inhibition were evaluated by analysis of the progress curves obtained with two procedures: the reaction initiated by addition of the enzyme and the reaction initiated by addition of the substrate after preincubation of the enzyme with the inhibitor. The obtained results were characteristic of slow-binding inhibition. The effects of different inhibitor concentrations on the initial and steady-state velocities obeyed the relationships of two-step enzyme-inhibitor interaction, qualified as mechanism B. It was found that TCoBQ and TCpBQ are strong urease inhibitors. TCpBQ is more effective than TCoBQ with the overall inhibition constant of K(i)* = 4.5 x 10(-7) mM. The respective inhibition constant of TCoBQ was equal to: K(i)* = 2.4 x 10(-6) mM. The protective experiment proved that the urease active site is involved in the tetrachlorobenzoquinone inhibition process. High effectiveness of thiol protectors against inhibition by TCoBQ and TCpBQ indicates the strategic role of the active site sulfhydryl group in the blocking process. The stability of the complexes: urease-TCoBQ and urease-TCpBQ was tested in two ways: by dilution or addition of dithiothreitol. No recovery of urease activity bound in the urease-inhibitor complexes proves that the complexes are stable and strong.

Antidepressant-like effect of lectin from Canavalia brasiliensis (ConBr) administered centrally in mice.

This study investigates the action of the central administration of the lectins isolated from Canavalia brasiliensis seeds (ConBr) and from Canavalia ensiformes seeds, (Concanavalin A, ConA) in the forced swimming test (FST) in mice. ConBr (1-10 micro g/site, i.c.v.), but not ConA, produced a decrease in the immobility time in the FST (observed at the time points 15, 30, 60 and 120 min after the injection), without changing the locomotor activity in the open-field test. The effect of ConBr in the FST was dependent on its protein structure integrity. ConBr (0.1 micro g/site, i.c.v.) caused a potentiation of the action of fluoxetine, a selective 5-HT reuptake inhibitor. The anti-immobility effect elicited by ConBr (10 micro g/site, i.c.v.) in the FST was prevented by the pretreatment of mice with pindolol (32 mg/kg, a 5-HT(1A/1B) receptor/beta-adrenoceptor antagonist), NAN-190 (0.5 mg/kg, a 5-HT(1A) receptor antagonist), ketanserin (5 mg/kg, a 5-HT(2A/2C) receptor antagonist), sulpiride (50 mg/kg, a D(2) receptor antagonist) or yohimbine (1 mg/kg, an alpha(2)-adrenoceptor antagonist), but not with SCH 23390 (0.05 mg/kg, a D(1) receptor antagonist) or prazosin (1 mg/kg, an alpha(1)-adrenoceptor antagonist). These results indicate that the antidepressant-like effect of ConBr in the FST is dependent on its interaction with the serotoninergic (via 5-HT(1A) and 5-HT(2)), noradrenergic (via alpha(2)-adrenoceptors) and dopaminergic (via D(2) receptors) systems. Considering the presence of lectins in the brain and based on the results, it will be important to determine a possible role of endogenous lectins in the modulation of the central nervous system function.