Purification, Characterization and Antimicrobial Properties of Hemolymph Lectin from the Larva of Red Palm Weevil, Rhynchophorus ferrugineus

Lectins are renowned hemagglutinins and multivalent proteins with a well-known quality for sugar-binding specificity that participate significantly in invertebrate defense functions. Studies on biological activity of lectin from coleopteran insect are very scarce. A lectin with specific affinity for N-acetyl neuraminic acid was purified from the hemolymph of the larva of the red palm weevil, Rhynchophorus ferrugineus by biospecific adsorption using formalinized rabbit erythrocytes and affinity chromatography using PSM-linked cyanogen bromide activated Sepharose 4B. The specific activity of the lectin purified by affinity chromatography was much higher than the lectin purified by biospecific adsorption. The binding specificity of the weevil lectin distinguishes it from other known insect lectins. Like the crude agglutinin, the lectin purified by affinity chromatography also showed the same pattern of specificity towards erythrocytes. However, 4-to-8-fold decrease in HA titer was observed when tested with the purified lectin. In the same way, reduction is also observed in the HAI titer of the purified lectin with most of the inhibitors except PSM where the HAI titer was identical both in the crude agglutinin and purified lectin. Sugars N-acetyl neuraminic acid, N-acetyl mannosamine and N-acetyl-D-galactosamine inhibited the HA titer of the purified lectin with greater efficacy than the crude agglutinin. The sialic acid specificity of the lectin was confirmed by 16-fold reduction in HA titer with asialo rabbit erythrocytes and 32-fold reduction in HAI titer with desialylated PSM. The purified lectin is homogenous on sodium dodecyl sulphate-polyacrylamide electrophorogram with a molecular weight of about 60 kDa. The lectin showed antimicrobial activity against pathogenic bacteria Streptococcus mutans, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeroginosa and fungi Candida albicans and Aspergillus niger

Physico-Chemical Characterization Of A Naturally Occurring Agglutinin In The Hemolymph Of The Larva Of Red Palm Weevil, Rhynchophorus Ferrugineus

The hemolymph of most invertebrates contains naturally occurring hemagglutinins with diverse specificities for erythrocytes. A natural agglutinin with high affinity for rabbit erythrocytes was detected in the hemolymph of the larva of the red palm weevil, Rhynchophorus  ferrugineus. The HA titer can be ranked as rabbit > guinea pig = dog = horse > human B > human O = buffalo > goat = pig > human A = rat = mice = cow erythrocytes.  Maximum hemagglutinin activity with rabbit erythrocytes was observed at pH 7.5 and temperature 30°C to 40°C.  The hemagglutinability was calcium dependent and reversibly sensitive to EDTA. The HA activity of the larval hemagglutinin was inhibited by the glycoproteins, Porcine stomach mucin = Fetuin > Bovine thyroglobulin > Transferrin = Apotransferrin > Bovine submaxillary mucin and sugars, N-acetyl mannosamine = melibiose = α-lactose > D-galactosamine > N-acetyl-D-galactosamine = N-acetyl neuraminic acid. Reduction in HA titer with the sialidase treated rabbit erythrocytes revealed the sialic acid specificity of the agglutinin. Presence of a single agglutinin and its proteinaceous nature were unveiled by cross adsorption studies and treatment with denaturing agents respectively.  Therefore, the preliminary description of the hemolymph agglutinin would offer ways to perform lectin purification from Rhynchophorus ferrugineus, the red palm weevil.&nbsp