A novel lectin from Sarcophaga. Its purification, characterization, and cDNA cloning

A novel C-type lectin that agglutinates rabbit red cells was purified from NIH-Sape-4 cells derived from the flesh fly (Sarcophaga peregrina), and its cDNA was isolated. This lectin, named granulocytin, appeared to be a trimer of a 20-kDa subunit consisting of 151 amino acid residues. The gene for granulocytin was activated in third instar larvae, and its expression was enhanced when the larval body wall was injured. In third instar larvae, granulocytin was found to be synthesized by hemocytes and secreted into the hemolymph. The molecular mass and gene expression patterns of granulocytin were very similar to those of Drosophila lectin that we reported previously (Haq, S., Kubo, T., Kurata, S., Kobayashi, A., and Natori, S. (1996) J. Biol. Chem. 271, 20213-20218). However, these two lectins showed amino acid identities of 20% at most, and no significant hapten sugar for granulocytin was identified.     

Purification, physicochemical characterization and biological properties of a lectin from Erythrina velutina forma aurantiaca seeds

A lectin was purified from seeds of Erythrina velutina forma aurantiaca by affinity chromatography on cross-linked guar gum. The lectin is a potent agglutinin for human (minimal concentration of protein able to cause visible agglutination of a 2% erythrocyte suspension varying from 1 to 4 micrograms/ml), rabbit (4 micrograms/ml) and chicken erythrocytes (8 micrograms/ml) but presented low activity against cow (250 micrograms/ml) or sheep (333 micrograms/ml) blood cells. Hemagglutination of human O+ erythrocytes was inhibited by D-lactose (0.2 mM) > D-galactose (0.8 mM) > D-raffinose (2.1 mM). At pH 7.5, chromatography on a Superose 12 HR 10/30 column showed that the lectin was primarily a dimer (56.0 kDa) composed of two identical subunits (31.6 kDa each). A small amount of a tetrameric form was also apparently present. The lectin is a glycoprotein (7.3% carbohydrate), has a pI of 4.5, contains high levels of acidic (Asp and Glu, 64.2 and 51.6 residues/mol, respectively) and hydroxy amino acids (Ser and Thr, 42.9 and 38.5 residues/mol, respectively) but relatively low amounts of sulfur amino acids (Cys and Met, 1.0 and 5.0 residues/mol, respectively) and has an N-terminal sequence of Val-Glu-Thr-Ile/Leu-Pro-Phe-Ser. Its hemagglutinating activity was abolished by heating at 70 degrees C for 10 min. The activation energy (delta G') required for denaturation measured by loss of hemagglutination activity was 24.87 kcal/mol. In rats, the purified lectin (100 micrograms) induced neutrophil migration into the peritoneal cavity (3.7 +/- 0.6 x 10(6) neutrophils/ml) or into the air pouch (2.75 +/- 0.25 x 10(6) neutrophils/ml), 8 and 10 times greater than the negative control, respectively.     

Tubulin structure: insights into microtubule properties and functions

The antimicrobial, hemagglutinating and toxic activities of the purple fluid of the sea hare Aplysia dactylomela are described. Intact or dialyzed purple fluid inhibited the growth of species of Gram-positive and Gram-negative bacteria and the action was not bactericidal but bacteriostatic. The active factor or factors were heat labile and sensitive to extreme pH values. The fluid preferentially agglutinated rabbit erythrocytes and, to a lesser extent, human blood cells, and this activity was inhibited by the glycoprotein fetuin, a fact suggesting the presence of a lectin. The fluid was also toxic to brine shrimp nauplii (LD50 141.25 micrograms protein/ml) and to mice injected intraperitoneally (LD50 201.8 +/- 8.6 mg protein/kg), in a dose-dependent fashion. These toxic activities were abolished when the fluid was heated. Taken together, the data suggest that the activities of the purple fluid are due primarily to substance(s) of a protein nature which may be involved in the chemical defense mechanism of this sea hare.     

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.     

Affinity purification of N-acetylglucosamine specific lectins. Purification and partial charactersation of triticale lectin

Seeds of Triticale contain a lectin that agglutinates rabbit erythrocytes whose activity is inhibited by N-Acetylglucosamine and its oligomers. An affinity method has been developed that efficiently binds the lectin activities from Triticale seeds and wheatgerm. Purified Triticale lectin is a glycoprotein with 3% carbohydrate and migrates as a single band corresponding to Mr. 15000 on SDS-PAGE. Antibodies raised to purified wheat germ agglutinin do not cross-react with purified Triticale lectin. Leucine/Isoleucine is the only terminal residue in the Triticale lectin. Presence of inhibitory sugar induces spectral changes in the protein in the UV region. Triticale lectin does not agglutinate human ABO erythrocytes and does not inhibit fungal growth.     

Characterization of a lectin from goat peripheral blood lymphocytes

A D-glucose specific lectin was isolated from goat peripheral blood lymphocytes by affinity chromatography on N-acetyl D-glucosamine agarose gel. The fluorescence intensity of 4 methyl umbelliferyl D-glucose was quenched to about 62% on addition of the lectin. This lectin gave a single band corresponding to 112 kDa in SDS-PAGE irrespective of treatment with 2-mercaptoethanol. The molecular weight and the Stoke's radius of the lectin in the native conditions were found to be 114 kDa and 4.54 nm, respectively, as determined by gel filtration on Sephacryl S 500 column. The lectin was found to be a glycoprotein with 5.6% of neutral hexose content and 5.5% of sialic acid. The lectin agglutinated trypsinized rabbit erythrocytes and human type A erythrocytes. The hemagglutinating activity was dependent on the presence of divalent cations like Mn2+ and Ca2+. Optimum pH, ionic strength and temperature for rebinding of lectin to acid treated Sephadex G200 were found to be 7.5, 0.16 and 30-37 degrees C, respectively.     

The lectin from the crustacean Liocarcinus depurator recognizes O-acetylsialic acids

A lectin that recognized sialic acids and aggultinated mouse erythrocytes was purified from hemolymph of the crab Liocarcinus depurator. It consisted of 38-kDa subunits and had a pI about 6.0. The specificity of the lectin was assayed by hemagglutination inhibition. N-acetylneuraminic acid (Neu5Ac) was a good inhibitor and its N-acetyl group at C-5 was critical for lectin-ligand interaction. Substitution of the C-9 hydroxyl on Neu5Ac with an O-acetyl group (9-O-Ac-Neu5Ac) increased the inhibitory potency of this molecule. Furthermore, O-acetyl substitution of all the hydroxyl groups yielded even better inhibitors (2,4,7,8,9-O-Ac-Neu5Ac and its 1-O-methyl ester). Removal of the hydroxyl or O-acetyl group connected to C-2 reduced the potency of these inhibitors. The lectin agglutinated and stimulated human but not mouse lymphocytes. It was also inhibited by Escherichia coli (O111:B4) lipopolysaccharide and agglutinated specific gram-negative bacteria. In vitro labeling with [35S]methionine indicated that the lectin was synthesized in hepatopangreas of L. depurator. Immunofluorescence showed that among hemocytes it localized mainly in the large-granule population.     

Lectin-induced apoptosis of tumour cells

The mechanisms of cytotoxic activity of Griffonia simplicifolia 1-B4 (GS1B4) and wheat germ agglutinin (WGA) lectins against various murine tumour cell lines were studied. Tumour cells that lack lectin-binding carbohydrates were resistant to lysis by these lectins. However, YAC-1 cells that expressed GS1B4 lectin-binding sites showed low sensitivity to lysis. To further analyse the relative importance of cell surface carbohydrates in lectin cytotoxicity, BL6-8 melanoma cells, which do not express the alpha 1,3 galactosyltransferase (alpha 1,3GT) gene and cell surface alpha-galactosyl epitopes reacting with GS1B4 lectin, were transfected with cDNA encoding alpha 1,3GT. After transfection, BL6-8 cells expressed high levels of GS1B4-binding alpha-galactosyl epitopes, but remained resistant to lysis by GS1B4 lectin, suggesting that the presence of lectin-binding epitopes, while essential, is not sufficient for tumour cell lysis and probably some intracellular mechanisms are involved in the regulation of lectin-mediated cytotoxicity. We found that the GS1B4 and WGA lectins induced apoptosis with DNA fragmentation of sensitive, but not resistant, tumour cell lines. DNA fragmentation, as well as tumour cell lysis, was blocked in the presence of the specific inhibitory sugar. To determine whether binding of the lectin to cell surface carbohydrates is sufficient to trigger tumour cell lysis, lectin-sensitive CL8-1 melanoma cells were incubated with GS1B4 lectin immobilized on agarose beads. Although these tumour cells bind to the immobilized lectin, it failed to trigger tumour cell death, suggesting that only soluble lectin is capable of tumour cell lysis and lectin internalization is probably required for their lysis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Tachycitin, a small granular component in horseshoe crab hemocytes, is an antimicrobial protein with chitin-binding activity

Small granules of horseshoe crab hemocytes contain two known major antimicrobial substances, tachyplesin and big defensin (S5), and at least five protein components (S1 to S6), with unknown functions. In the present study, we examined the biological properties and primary structure of a small granular component S2, named tachycitin. This component was purified from the acid extract of hemocyte debris by two steps of chromatography. The purified tachycitin was a single chain protein with an apparent M(r) = 8,500 on Tricine-SDS-polyacrylamide gel electrophoresis. Ultracentrifugation analysis revealed tachycitin to be present in monomer form in solution. Tachycitin inhibited the growth of both Gram-negative and -positive bacteria, and fungi, with a bacterial agglutinating property. Moreover, tachycitin and big defensin acted synergistically in antimicrobial activities. The amino acid sequence and intrachain disulfide bonds of tachycitin were determined by amino acid and sequence analyses of peptides produced by enzymatic cleavages. The mature tachycitin consisted of 73 amino acid residues containing five disulfide bonds with no N-linked sugar. A cDNA coding for tachycitin was isolated from a hemocyte cDNA library. The open reading frame coded for an NH2-terminal signal sequence followed by the mature peptide and an extension sequence of -Gly-Arg-Lys at the COOH-terminus, which is a putative amidating signal. The COOH-terminal threonine amide released after digestion of tachycitin with lysylendopeptidase was identified. The NH2-terminal 28 residues of tachycitin shows sequence homology to a part of chitin-binding regions found in antifungal chitin-binding peptides, chitin-binding lectins, and chitinases, all of which have been isolated from plants. Tachycitin showed a specific binding to chitin but did not bind with the polysaccharides cellulose, mannan, xylan, and laminarin. Tachycitin may represent a new class of chitin-binding protein family in animals.     

Purification and characterization of a mannose/glucose-specific lectin from Castanea crenata

A hemagglutinin was purified from the cotyledons of Japanese chestnut (Castanea crenata Sieb. et Zucc.) by affinity chromatography on asialo-fetuin Sepharose 4B column followed by anion-exchange and gel permeation chromatography. The hemagglutinating activity of Castenea crenate agglutinin (CCA) was strong for sialidase-treated human erythrocytes, but was inhibited by mannose, glucose, and their derivatives as well as by glycoproteins having an N-linked complex carbohydrate type. The apparent M(r) of intact CCA was determined to be ca 257,000 by gel filtration using a Superose 12 column. In SDS-PAGE, under reducing and non-reducing conditions, CCA migrated as a single band of M(r) 37,000. Therefore, the intact CCA may be composed of six or eight identical subunits without disulfide bonds. In addition, CCA showed strong mitogenic activity similar to other lectins. The N-terminal amino acid of CCA may be blocked since no amino acid was detected by direct sequence analysis. Amino acid analysis showed that CCA was rich in glycine, but did not contain cysteine residues. Some properties of CCA were similar to mannose/glucose-specific legume lectins, but our data suggest that the molecular structure of CCA is different.     

Elucidation of lectin receptors by quantitative inhibition of lectin binding to human erythrocytes and lymphocytes

The binding to normal and sialidase-treated human erythrocytes and lymphocytes of four 125I-labeled lectins [Maackia amurensis hemagglutinins (MAM and MAH), Ricinus communis hemagglutinin (RCH), and Bauhinia purpurea hemagglutinin (BPH)] was studied in detail. The quantitative inhibition assays against the lectin binding to the cells were also performed with various glyco-proteins and glycopeptides as inhibitors. The comparison of the inhibition constants of the inhibitors thus obtained with the association constants of the lectins to the cells permitted estimation of the relative receptor activities of cell surface glyco-proteins toward the lectins.     

Hemagglutinating activity of extracellular alkaline metalloendopeptidases from Vibrio sp. NUF-BPP1

Alkaline metalloendopeptidase (metalloprotease) AP1 (48 kDa) from Vibrio sp. isolated from the intestine of a five-barred goatfish (Parupeneus trifasciatus) was reported in our previous paper to produce AP2 (36 kDa) by releasing a peptide fragment (molecular mass of about 12 kDa) from the C-terminal end of AP1 by autodigestion. AP1 strongly agglutinated fish (flounder, Paralichthys olivaceus) and rabbit erythrocytes, and weakly chicken erythrocytes. In contrast, AP2 had no significant hemagglutinating activity toward any erythrocytes tested, except for weak activity on flounder erythrocytes, suggesting that the C-terminal region of AP1 may be required for the strong hemagglutinating activity. The optimum temperature for the hemagglutinating activity of AP1 was found to be lower than that for the proteolytic activity. At acidic pHs (below pH 7.5), the hemagglutinating activity of AP1 decreased, and its pH profile resembled that of the proteolytic activity. The hemagglutinating activity of AP1 was not observed in the presence of o-phenanthroline or synthetic and proteinous substrates, but different kinds of saccharides and lipids had no effect. While the proteolytic activity of AP1 was not affected by CaCl2, the hemagglutinating activity of AP1 decreased with increases in CaCl2 concentrations. These results suggested that the hemagglutinating activity of these proteases (AP1 and AP2) was most likely caused by their proteolytic action on erythrocyte cell surfaces.     

Korormicin, a novel antibiotic specifically active against marine gram-negative bacteria, produced by a marine bacterium

A novel antibiotic named korormicin was isolated from the marine bacterium, Pseudoalteromonas sp. F-420. This strain was isolated from the surface of a macro alga Halimeda sp. collected from Palau (the Republic of Belau). The planar structure of korormicin was determined by the result of 2D NMR studies and mass spectral data. Korormicin had specific inhibitory activity against marine Gram-negative bacteria, but was inactive against terrestrial microorganisms.     

Vibrio mimicus attaches to the intestinal mucosa by outer membrane hemagglutinins specific to polypeptide moieties of glycoproteins

Vibrio mimicus is the closest organism to Vibrio cholerae. V. mimicus E-33, which is a highly adhesive and enteropathogenic strain, is known to produce three types of hemagglutinins (HAs), i.e., a 31-kDa exocellular metalloprotease (Vm-HA/protease), lipopolysaccharide (Vm-LPSHA), and a 39-kDa major outer membrane protein (Vm-OMPHA). Hemagglutination induced by Vm-LPSHA and Vm-OMPHA was inhibited by glycoproteins, including mucin, fetuin, and asialofetuin, but not by monosaccharides, disaccharides, or N-acetylated saccharides. The inhibitory potential of each glycoprotein for Vm-OMPHA was greatly augmented by treatment with a glycolytic enzyme such as beta-D-galactosidase or beta-D-glucosidase, while pronase treatment achieved complete abolition of the inhibitory potential. The inhibitory ability of the glycoproteins for Vm-LPSHA was also abolished by pronase treatment; however, glycolytic enzyme treatment showed no effect. Hence, the polypeptide portion of glycoproteins may directly associate with Vm-OMPHA and Vm-LPSHA, but the sugar moiety may act as a barrier to interaction with Vm-OMPHA. The glycoproteins as well as Fab antibodies against Vm-OMPHA and Vm-LPSHA eliminated the ability of E-33 cells to agglutinate rabbit erythrocytes and to attach to rabbit intestinal mucosa. Additionally, expression of the hemagglutinating ability by the bacterial cells was accompanied by efficient bacterial adherence to the intestinal mucosa. Finally, the hemagglutinating activity of Vm-OMPHA was markedly increased by incubation with Vm-HA/protease. These results indicate that all three HAs may have significant roles in the glycoprotein-mediated intestinal adherence of V. mimicus E-33.     

Separation of bacterial capsular and lipopolysaccharides by preparative electrophoresis

Preparative gel electrophoresis was used to separate and purify extracellular, capsular and lipopolysaccharides (EPSs, CPSs, and LPSs, respectively) from crude bacterial extracts. The procedure effectively separates CPS from LPSs. In addition discreet size ranges of these various polysaccharides can be isolated. The 'rough' (R-type), 'smooth' (S-type), and 'semi-smooth' LPSs were separated from one another. In addition different size classes of 'semi-smooth', or S-type LPS, can be separated. This procedure was demonstrated for diverse bacterial species, including the soil bacteria Rhizobium fredii, and the enteric bacterial species, Salmonella enteritidis and Proteus mirabilis. In the latter case, it was also possible to separate capsular polysaccharide from its lipid-bound form.     

Purification and characterization of the agglutinins from the sponge Aaptos papillata and a study of their combining sites

The lectins from the sponge Aaptos papillata were isolated by affinity chromatography using polyleucyl blood group A + H substances from hog stomach linings as an absorbent and eluting with 3 M MgCl2. Further separation on diethylaminoethylcellulose and preparative disc electrophoresis on polyacrylamide gave the three fractions, Aaptos lectins I, II, and III. They were essentially homogenous in polyacrylamide electrophoresis and sedimentation analysis: a small second component was seen in lectins I and II in immunoelectrophoresis at high concentration. The S20,W0 values for Aaptos lectins I, II, and III were 3.5, 6.0, and 5.5. By electrophoresis in sodium dodecyl sulfate with an without beta-mercaptoethanol Aaptos lectin I showed two bands corresponding to molecular weights of 12 000 and 21 000; Aaptos lectins II and III gave only one band of molecular weight of 16 000. In isoelectric focusing, Aaptos lectin I showed bands at pH 4.7 and 5.4 and in the range between 6.8 and 7.6, while Aaptos lectins II and III were almost identical with bands at pH 3.8, 4.7 to 4.9, and 5.3. Aaptos lectin I differed from II and III in amino acid composition but the latter two were very similar. They contained no significant carbohydrate. Aaptos lectin I reacted best with blood group substances with terminal nonreducing N-acetyl-D-glucosamine residues precipitating about two-thirds of the lectin N added while blood group substances with terminal nonreducing DGalNAc were almost inactive. However, Aaptos lectin II was completely precipitated by blood group substances and glycoproteins containing terminal DGalNAc, DGlcNAc, or sialic acid residues. Aaptos lectin III had a precipitation pattern similar to Aaptos lectin II. DGlcNAc but not DGalNAc inhibited precipitation of Aaptos lectin I by blood group substances and N, N', N', N'-tetraacetylchitotetraose was the best inhibitor and was 2000 times more active than DGlcNAc. Precipitin reactions with Aaptos lectin II were inhibited by equal amounts of DGlcNAc and by sialic acid which were four times more potent than DGalNAc. N,N',N'-triacetylchiotriose was the best inhibitor and was 13 times better than DGlcNAc. At 37 degrees C three to four times higher amounts of inhibitor were necessary to inhibit precipitation of Aaptos lectin II than were needed at 4 degrees C, indicating higher affinity of blood group substances for Aaptos lectin II with increasing temperature. Aaptos lectin I was precipitated by the monofunctional hapten p-nitrophenyl-alphaDGalNAc, while p-nitrophenyl-betaDGalNAc did not precipitate and was a good inhibitor. Both phenomena indicate involvement of hydrophobic bonds.     

Characterization and representative structures of N-oligosaccharides bound to apolipoprotein H

We studied the structure of N-linked carbohydrates bound to apolipoprotein H by a combination of two methods which make use of lectins. Digoxigenin-labelled lectins are used for the structural characterization of carbohydrate chains of glycoproteins. Concanavalin A lectin affinity chromatography was used to analyse apolipoprotein H according to the characteristics of its carbohydrate chain inner to sialic acid residues. Our results from digoxigenin-labelled lectins analysis showed that apolipoprotein H gave positive bands to SNA, DSA, GNA, PNA and AAA lectins. Apolipoprotein H gave a negative band when reacted with MAA lectin. When we applied apolipoprotein H onto the Concanavalin A lectin column no detectable amounts of protein were eluted with Concanavalin A buffer. After adding a buffer with low sugar concentration (10 mM glucoside) a large amount of apolipoprotein H was recovered. These molecules of apolipoprotein H weakly bound to the lectin. When a higher sugar concentration (500 mM mannoside) was added most of the sample applied was eluted. These molecules of apolipoprotein H firmly bound to the column having high affinity for the lectin. These results combined with those coming from the digoxigen-labeled lectins method enable us to understand the inner structure of carbohydrate chains with their outer branches. Molecules of apolipoprotein H which weakly bind to Concanavalin A could bear complex N-glycans organized in biantennary or truncated hybrid structures. Firmly bound apolipoprotein H referred to molecules rich in N-glycan hybrid structures. They have an outer branch belonging to the high mannose carbohydrate chains which explain the ability to bind to the column and an other main branch bearing the sequence galactose beta-(1-4)-N-acetylglucosamine beta-(1-2) mannose. Galactose could be the terminal sugar or, alternatively, be masked with sialic acid alpha-(2-6) terminally linked.     

Macromolecular recognition: effect of multivalency in the inhibition of binding of yeast mannan to concanavalin A and pea lectins by mannosylated dendrimers

The synthesis and binding properties of a new family of high affinity alpha-D-mannopyranoside ligands are described. The synthesis of the new multivalent ligands is based on the scaffolding of multiantennary branches of L-lysine residues having electrophilic N-chloroacetylated end groups as core structures. An alpha-D-mannopyranoside with p-substituted aryl aglycon ending with a thiol group was prepared and covalently attached to each of the branches of the dendritic structures. The resulting glycodendrimers with 2 (12), 4 (14), 8 (16), and 16 (18) mannoside residues were tested for their relative inhibitory potency by solid-phase enzyme-linked lectin assays (ELLA) using methyl and p-nitrophenyl alpha-D-mannopyranosides as standards. Concentrations necessary for 50% inhibition (IC50s) of binding of yeast mannan to Jack bean phytohemagglutinin (Canavalia ensiformis, concanavalin A) and to pea lectin (Pisum sativum) were determined. Analogous mannosylated copolyacrylamides were also prepared for comparison. The IC50 values were also plotted as a function of dendrimer valencies. The inhibitions showed 16-mer 18 to be approximately 600- and 2000-fold more potent than methyl alpha-D-mannopyranoside, and 66- and 1383-fold more potent than p-nitrophenyl alpha-D-mannopyranosides with Con A and pea lectins, respectively. Even when these numbers are expressed relative to single mannopyranoside residues per dendrimers, the relative potencies against the aromatic mannoside are still 4- and 86-fold better against Con A and pea lectins. These results unequivocally indicate that the optimum inhibitory binding properties of the new mannosylated dendrimers vary with both dendrimers and lectin valencies.     

Characterization and molecular cloning of the lectin from Helianthus tuberosus

A lectin called Helianthus tuberosus agglutinin or Heltuba has been isolated from tubers of the Jerusalem artichoke, a typical representative of the Asteraceae family. Heltuba is a tetrameric protein composed of four identical subunits of 15.5 kDa and exhibits a preferential specificity towards oligomannosides. Cloning of the corresponding cDNAs revealed that the mature lectin polypeptide comprises the entire open reading frame of the cDNA suggesting that the primary translation product is not processed and that the lectin is a cytosolic protein. Searches in the databases revealed sequence similarity with lectins from the taxonomically unrelated Convolvulaceae and Moraceae species. Therefore, the discovery of Heltuba is of great importance in view of the occurrence and molecular evolution of the jacalin-related lectins.