**8. Structure of plant lectin**

154 Antimicrobial Agents

As mentioned, lectin interacts with galactose or *N*-acetylgalactosamine such as the lectin from the corn coleoptyle. It is a glycoprotein had molecular mass under non-denaturing conditions was 88.7 kDa And had carbohydrates that constituted 12% of the total weight comprised galactose, mannose, and *N*-acetyl-D-glucosamine (Martinez-Cruz et al., 2001). In 2003 Konozy et al. were found *Erythrina speciosa* seeds can be specific with D-galactose and had two identical subunits of molecular mass was 27.6 kDa include the lectin was a neutral carbohydrate content of 5.5% (Konozy et al., 2003). *N*-acetyl-D-galactosamine-specific lectin isolation from *Glycine max* L. Merrill SA88 them were found the soybean lectin consists of four subunits it had molecular weight of each 30,000 Da in one-step purification with high purity and high yield (about 90% recovery from the crude extract) by use Poly (hydroxypropyl methacrylate-glycidyl methacrylate) beads were as an affinity matrix and

*N*-acetyl-D-galactosamine (GalNAc) was as an affinity ligand (Percin et al., 2009).

*Aleuria aurantia* lectin (AAL) is a commercially available lectin that is known for its high affinity for α-1, 6-fucosylated oligosaccharides and it is widely used to estimate the extent of α-1,6-fucosylation on glycoproteins and to fractionate glycoproteins. For research a novel probe for core fucose from *Aspergillus oryzae* L-fucose-specific lectin (AOL) has strongest preference for the alpha 1,6-fucosylated chain among α-1,2-, α-1,3-, α-1,4-, and α-1,6 fucosylated pyridylaminated (PA)-sugar chains. These results suggest that AOL is a novel probe for detecting core fucose in glycoproteins on the surface of animal cells (Matsumura et al., 2007). Furthermore, *Lotus tetragonolobus* lectin is a fucose-specific legume lectin. It is a homotetramer composed of four legume lectin domains was 27,800 Da (Moreno et al.,

Most of Sialic acid-specific lectins was found in invertebrates such as those from the Indian horseshoe crab (Mohan et al., 1982), marine crab *Scylla serrata* (Mercy and Ravindranath, 1992), lobster, tunicalase, fungus *Hericium arinaceum* (Kawagishi et al., 1994) and leaves of mulberry (Ratanapo et al., 1998). A lectin from the white shrimp *Litopenaeus setiferus* (LsL) hemolymph is a heterotetramer of two 80 kDa and two 52 kDa subunits, *N*-acetylated sugars, such as GlcNAc, GaINAc, and NeuAc, were the most effective inhibitors of the LsL hemagglutinating activity. Desialylation of erythrocytes or inhibitory glycoproteins abolished their capacity to bind LsL, confirming the relevance of sialic acid in LsL-ligand interactions (Alpuche et al., 2005). In 2009 the *Phaseolus coccineus* lectin (PCL) specificity towards sialic acid showed the molecular mass of 30 kDa consisting of homodimer subunits. Moreover the purified PCL was devoid of antifungal activity against *Candida albicans* and Penicillium *italicum*, but markedly inhibited the growth of *Hericium maydis*, *Rhizoctonia solani*, Gibberella *sanbinetti,* and Sclerotinia *sclerotiorum* while the same concentration of PCL decrease the 50% hemagglutinating activity was inhibited by sialic acid it suggesting a significant correlation between sialic acid-specific site and its bi-functional bioactivities

**7.2 Galactose/***N***-acetylgalactosamine** 

**7.3 Fucose** 

2008).

**7.4 Sialic acids** 

(Chen et al., 2009).

Different lectin families are in general structurally unrelated. And even in those cases where a common fold is recruited, convergent evolution is the most likely explanation. Some lectin families such as the galectins recognize only one specific oligosaccharide, and consequently have a very conserved recognition site. On the other extreme, members of the C type lectin family span a wide variety of specificities. Consequently, their recognition sites are highly variable, and different specificities can easily be engineered by site directed mutagenesis (Iobst and Drickamer, 1994; and Kolatkar and Weiss, 2009). A general feature of binding sites of all lectins seems to be that they consist of a primary binding site that is capable of recognizing in a specific way a single monosaccharide residue, usually with a low affinity (in the millimolar range). Very often, but not always, there are further subsites that can be occupied by sugar residues connected to the one bound in the primary site. This allows for a modest increase in affinity. Folding in common between plant and animal lectins are βsandwich fold, β-Trefoil folds and Hevein domains. The legume lectin-like β-sandwich fold found in Galectins that conserved family of β-galactosyl binding lectins that occur in both vertebrates and invertebrates (Hirabayashi et al., 2009). Except for the legume lectins, galectins and pentraxins, it is observed in a number of carbohydrate processing and other enzymes such as β-glucanase and asparagine amidase (Keitel et al., 1993; and Kuhn et al., 1994).

Fig. 3. The legume lectin-like β-sandwich folds illustrated by a member of the legume lectins (Concanavalin A in complex with the trisaccharide [Man(α-1, 3)]Man(α-1, 6)Man). In each case, only a single monomer of the multimeric protein is shown. Bound carbohydrate is shown in ball-and-stick. Metal ions are shown as grey spheres, bound ligands are shown in green ball-and-stick representation (Loris, 2002).

The β-trefoil fold was first identified as a carbohydrate recognition domain in ricin (Montfort et al., 1987). Later, it was also found to be the fold of amaranthin. The β-trefoil fold is another fairly common fold, first identified in soybean trypsin inhibitor (Sweet et al. 1974). It consists of a repeat of three subdomains, each consisting of a fourstranded antiparallel β-sheet.

Antimicrobial Activity of Lectins from Plants 157

There are no structural features common to all lectins. Many of these proteins are relatively rich in aspartic acid, serine and threonine, which comprise as much as 30% of their amino acid content and are low in sulfur-containing amino acids. Such a pattern of amino acids is characteristic of plant proteins. In contrast, lectins such as those from wheat germ, potato and pokeweed are rich in cysteine with 20, 11.5 and 18% of the total amino acid residues respectively, most or all of which are in the form of cysteine. The high content of disulfide bonds in wheat germ agglutinin endows the protein with stability to heat (Aub et al., 1963), to proteolytic enzymes and to denaturing agents such as detergents, urea, alkali and acids (Nagata and Burger, 1972; and Rice and Etzler, 1974). The potato and *Datura stramonium*  lectins are rich in hydroxyproline (Lamport, 1969). A few lectins, such as Concanavalin A, wheat germ and peanut agglutinins are devoid of covalently bound sugars. Most lectins, however, are glycoproteins with carbohydrate contents that can be as high as 50%, e.g., potato lectin. The table shown below (Table 1) is on the sugar contents of certain important glycoprotein lectins. The sugar constituents in animal glycoproteins are the same as those

**Lectin Mannose Galactose L-Fucose L-Arabinose GlcNAc Xylose C-P linkage Reference** 

*Glycine max* 4.5 1.2 GlcNAc-Asn Lis et al.,

3 47 Ara-Hyp,

The molecular weight of lectins in plants ranges from 36,000 Da for wheat germ agglutinin (Nagata and Burger, 1972; and Rice and Etzler, 1974) to 265,000 for lima bean lectin (Galbraith and Goldstein, 1970). The lower limit of MW of animal lectins is found to be 14 kDa (Lis and Sharon, 1998). Some lectins exhibit a pronounced tendency to aggregate. Thus, the MW of Concanavalin A at pH below 6 is 51,000 Da and at physiological pH it is 12,000 Da (Mc Cubbin and Kay, 1971; and Wang et al., 1971). Upon storage at room temperature, soybean agglutinin and peanut agglutinin also possibly undergo irreversible self-association to high molecular weight aggregates (Lotan et al., 1975). The subunits are identical in most

5.8 1 2.6 1 Lescar et al.,

2.8 4.5 Kilpatrick,

3.2 3.7 0.5 1.3 GlcNAc-Asn Mach et al.,

7.3 2.8 GlcNAc-Asn Ohtani, K.,

0.77 1.63 0.65 GlcNAc-Asn Kurokawa

2007

1978

1973

1991

Gal-ser

and Misaki, A. (1980)

Matsumoto et al., 1983

et al., 1976

**9. Physicochemical properties of plant lectin** 

found in other plant glycoproteins, with the exception of L-arabinose.

**9.1 Composition** 

*Bandeiraea simplicifolia* 

*Datura stramonium* 

*Phaseolus lunatus* 

*Phaseolus vulgaris* 

*Solanum tuberosum* 

*Wistaria floribunda* 

Table 1. Well-characterized glycoprotein lectins

Fig. 4. The β-trefoil folds illustrated by the first domain of the subunit ricin (in complex with lactose), the first domain of amaranthin (in complex with the T-antigen) In each case, for clarity, only a single β-trefoil domain is shown in an identical orientation, although each protein is a multidomain as well as a multimeric protein. Bound carbohydrate is shown in green ball-and-stick (Loris, 2002).

Lectins comprise a structurally very diverse class of proteins characterized by their ability to bind carbohydrates with considerable specificity. Although lectins bind monosaccharides rather weakly, they employ common strategies for enhancing both the affinity and specificity of their interactions for more complex carbohydrate ligands. Members of the legume lectin family show considerable sequence and structural homology, but differences in their carbohydrate-binding specificity. The legume lectin monomer has a molecular weight of 25,000 and is composed primarily of a six- and a seven-stranded antiparallel-sheet. Concanavalin A (Con A), *Lathyrus ochrus* isolectin I (LOL I), and pea lectin all show mannose and glucose-binding specificity, and the X-ray crystal structures of their carbohydrate complexes show a monosaccharide binding- site geometry very similar to that of the LOL Iα-methyl-o-mannopyranoside complex.

The first lectin structures to be determined derived from two phylogenetically conserved families, the *Leguminoseae* (Sharon and Lis 2001; and Young and Oomen, 1992) and the *Gramineae* (Raikhel et al., 1993). Over the past ten years, major advances in X-ray crystallographic technology and the relative ease of isolation and crystallization of plant lectins have led to a rapid increase in crystal structures. The leguminous lectins clearly have dominated the field with some ten structures known today. Four of these, peanut lectin, soybean lectin, lentil lectin and hemagglutinin L, have been determined within the past three years. These lectins display diverse sugar-binding specificitiesistructures.

Lectins of the *Amaryllidaceae, Orchidaceae, Alliaceae, Araceae* and *Liliaceae* families are in the class of the mannosespecific *Liliatae* and constitute the third major structurally characterized plant lectin superfamily (Van Damme et al., 1994). They are nonseed lectins of multigene families isolated from plant bulbs (Van Damme et al., 1994) and function either as dimers or tetramers, as do the legume lectins. Their strict and exclusive specificity solely for mannose has imparted some unusual biological properties *in vitro* to this lectin family including their antiviral properties against retroviruses (e.g. HIV) (Hammaar et al., 2006). Snowdrop lectin (GNA) is a tetrameric lectin (Mr =50,000 Da) and is the first member of the *Amaryllidaceae*  family crystallographically investigated.
