¹H‐NMR structural determination of the N‐linked carbohydrate chains on glycopeptides obtained from the bean lectin phytohemagglutinin

Abstract

Phytohemagglutinin, the lectin of the common bean Phaseolus vulgaris, is a N-linked glycoprotein with one high-mannose-type and one xylose-containing oligosaccharide side chain per polypeptide. The high-mannose-type glycan is attached to Asnl2 and the complex-type glycan to Am60 [Sturm, A. & Chrispeels, M. J. (1986) Plant Physiol. 81, 320-3221. The structures of the oligosaccharides were elucidated from two glycopeptides obtained from the lectin by Pronase digestion, affinity chromatography on concanavalin-A -Sepharose and gel-filtration chromatography on a column of BioGel P-4. The N-linked glycan structures were investigated by 500-MHz 'H-NMR spectroscopy and were established to be:M m a ( l-Z)Mana( 1-6) D3

Core 1,3-fucose is a key part of the epitope recognized by antibodies reacting against plant N-linked oligosaccharides and is present in a wide variety of plant extracts

“…What maybe true of antibodies raised in rabbits could also be true of IgE pools from patients allergic to plant materials and if patients have IgE against the glycan, then cross-reactions not dependent on known conserved proteins are possible. Indeed, the xylosylated and/or fucosylated N-linked carbohydrate structures in plants are probably more conserved across plant species than any protein sequence, as shown by the studies referred to above (van Kuik et al, 1986;Ashford et al, 1987Ashford et al, , 1991Fournet et al, 1987;Sturm et al, 1987Sturm et al, , 1992D'Andrea et al, 1988;Balshüsemann and Jaenicke, 1990;Hayashi et al, 1990;Takahashi et al, 1990;Ramirez-Soto and Poretz, 1991;Sturm, 1991;Debray et al, 1992;Stahl et al, 1994;Gray et al, 1996;Ogawa et al, 1996;Ohsuga et al, 1996;Oxley et al, 1996;Yang et al, 1996;Altmann, 1998). Also, xylosylated N-glycans are present in snails (van Kuik et al, 1985), while α1,3-linked fucosylated N-glycans are present in insects (Kubelka et al, , 1995.…”

“…Cry j I contains elaborate glycans with core xylose and α1,3-linked fucose, BG60 allergen contains core α1,3-linked fucose (but no xylose) and the PNGase F released glycans of Art v II are all of the oligomannose type. That α1,3-fucose and β1,2-xylose residues are more common in plants than just on the glycoproteins against which antisera have been prepared is evidenced by many studies such as those on legume lectins (Ashford et al, 1991), pineapple stem bromelain (van Kuik et al, 1986), bean phytohemagglutinin (Sturm et al, 1992), bean phaseolin (Sturm et al, 1987), mistletoe lectin (Debray et al, 1992), miraculin , red kidney bean purple acid phosphatase (Stahl et al, 1994), soybean peroxidase , zucchini ascorbate oxidase (Altmann, 1998), and Sophora japonica lectin (Fournet et al, 1987). β1,2-Xylosylated oligosaccharides lacking α1,3-linked fucose have been found in rice amylase oligosaccharides (Hayashi et al, 1990), zucchini ascorbate oxidase oligosaccharides (D'Andrea et al, 1988;Altmann, 1998), algal oligosaccharides (Balshüsemann and Jaenicke, 1990), and self-incompatibility ribonucleases of Nicotiana alata (Oxley et al, 1996), as well as on snail hemocyanin oligosaccharides (with core α1,6-fucose rather than α1,3-fucose; van Kuik et al, 1985).…”

Distribution of xylosylation and fucosylation in the plant Golgi apparatus

“…As soon as they arrive in the trans Golgi cisternae, the glycoprotein N-linked complex glycans may receive an a l + 3 fucose on the proximal GlcNAc of the core. The glycan structure G~c N A c~M~~~X~~F U C ( G I C N A C )~ is then further matured either by removal of the two terminal GlcNAc in the vacuole, yielding to the PHA-type glycan structure Man,Xyl-FUC(GICNAC)~ (Sturm et a/., 1992;Vitale and Chrispeels, 1984;Vitale et a/., 1984) or by addition of al-16 fucose and P1+4 galactose residues, leading to the laccasetype glycan Fuc2Ga12GlcNAc2Man3XylFuc(GlcNAc)2 (Takahashi etal., 1986). This model describing a compartmental organization of complex glycan processing in the plant Golgi does not necessarily imply that glycosidase and glycosyltransferase activities have a very strict compartmentation over the secretory pathway.…”

Structure-Function Relationship of Monocot Mannose-Binding Lectins

“…In fetuin, the exposed Gal residues of both O-linked and N-linked saccharides are masked by sialic acid residues. PHA-E contains an N-linked highMan-type glycan with exposed Manal+Z-linked residues and a Xyl-containing oligosaccharide with a Fucal+3 branched residue (Sturm et al, 1992) (Fig. 1).…”