The glycosylation of glycoprotein lectins. Intra- and intergenus variation in N-linked oligosaccharide expression (original) (raw)
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How a Plant Lectin Recognizes High Mannose Oligosaccharides
PLANT PHYSIOLOGY, 2007
The crystal structure of Pterocarpus angolensis seed lectin is presented in complex with a series of high mannose (Man) oligosaccharides ranging from Man-5 to Man-9. Despite that several of the nine Man residues of Man-9 have the potential to bind in the monosaccharide-binding site, all oligomannoses are bound in the same unique way, employing the tetrasaccharide sequence Mana(1-2)Mana(1-6)[Mana(1-3)]Mana(1-. Isothermal titration calorimetry titration experiments using Man-5, Man-9, and the Man-9-containing glycoprotein soybean (Glycine max) agglutinin as ligands confirm the monovalence of Man-9 and show a 4-times higher affinity for Man-9 when it is presented to P. angolensis seed lectin in a glycoprotein context.
European Journal of Biochemistry, 2005
Glycopeptides and oligosaccharides of either the N-acetyllactosaminic or the oligomannosidic type derived from glycoproteins containing the N-glycosylamine linkage were used to define the specificity of different lectins (concanavalin A, Lens culinaris agglutinin, Vicia faba agglutinin, Pisum sativum agglutinin, Ricinus communis agglutinins, soybean agglutinin, wheat germ agglutinin, Solanum tuberosum agglutinin, Datura stramonium agglutinin, Lotus tetragonolobus agglutinin, Ulex europeus agglutinin) by studying the inhibition of human red blood cell agglutination by these structures. The results obtained show that lectins considered 'identical' in terms of monosaccharide specificity, possess the ability to recognize fine differences in more complex structures. In fact, different lectins are able to recognize different saccharidic sequences on the same glycan structure. As these sequences are likely to be common to numerous glycoproteins, including cell membrane glycoproteins, the results obtained with lectins in the study of cell surface carbohydrates have to be very carefully interpreted. Moreover, our results confirm previous data on the spatial configuration of the glycan moiety of glycoproteins deduced from the construction of molecular models: the fact that oligosaccharides bearing an alpha-NeuAc-(2 leads to 6)-Gal unit are more powerful inhibitors than oligosaccharides bearing an alpha-NeuAc-(2 leads to 3)-Gal unit could be related to the high rotational freedom of alpha-2,6 linkage; the observation that glycoasparagines, glycopeptides and glycoproteins possess a higher affinity for lectins than the related oligosaccharides could be explained by the fact that the glycan--amino acid linkage leads to structures more rigid than those of the oligosaccharides themselves.
Plant Physiology, 1986
ABSTRACI Phytohemagglutinin, the lectin of the common bean Phaseolus vulgaris, has a high mannose and a modified (fucosylated) oligosaccharide on each polypeptide. Fractionation by high performance liquid chromatography of tryptic digests of VHlfucose or [3H1glucosamine labeled phytohemagglutinin, followed by amino acid sequencing of the isolated glycopeptides, shows that the high mannose oligosaccharide is attached to Asn"2 and the modified oligosaccharide to Asn"' of the protein. In animal glycoproteins, high mannose chains are rarely found at the N-terminal side of complex chains.
Structure, 1994
Background: Lectins mediate cell-cell interactions by specifically recognizing oligosaccharide chains. Legume lectins serve as mediators for the symbiotic interactions between plants and nitrogen-fixing microorganisms, an important process in the nitrogen cycle. Lectins from the Viciae tribe have a high affinity for the fucosylated biantennary N-acetyllactosamine-type glycans which are to be found in the majority of N-glycosylproteins. While the structures of several lectins complexed with incomplete oligosaccharides have been solved, no previous structure has included the complete glycoprotein. Results: We have determined the crystal structures of Lathyrus ochrus isolectin II complexed with the N2 monoglycosylated fragment of human lactotransferrin (18kDa) and an isolated glycopeptide (2.1 kDa) fragment of human lactotransferrin (at 3.3A and 2.8A resolution, respectively). Comparison between the two structures showed that the protein part of the glycoprotein has little influence on either the stabilization of the complex or the sugar conformation. In both cases the oligosaccharide adopts the same extended conformation. Besides the essential mannose moiety of the monosaccharide-binding site, the fucose-l' of the core has a large surface of interaction with the lectin. This oligosaccharide conformation differs substantially from that seen in the previously determined isolectin I-octasaccharide complex. Comparison of our structure with that of concanavalin A (ConA) suggests that the ConA binding site cannot accommodate this fucose. Conclusions: Our results explain the observation that Viciae lectins have a higher affinity for fucosylated oligosaccharides than for unfucosylated ones, whereas the affinity of ConA for these types of oligosaccharides is similar. This explanation is testable by mutagenesis experiments. Our structure shows a large complementary surface area between the oligosaccharide and the lectin, in contrast with the recently determined structure of a complex between the carbohydrate recognition domain of a C-type mammalian lectin and an oligomannoside, where only the non-reducing terminal mannose residue interacts with the lectin.
Journal of Biological …, 1986
A lectin specific for chito-oligosaccharides from the exudate of ridge gourd (Luffa acutangula) fruits has been purified to homogeneity by affinity chromatography. The lectin has a molecular weight of 48,000, an SX,,,~ of 4.06 S and a Stokes radius of 2.9 nm. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a single band corresponding to M, of 24,000 was observed both in the presence and absence of &mercaptoethanol. The subunits in this dimeric lectin are, therefore, held together solely by noncovalent interactions. The lectin is not a glycoprotein, and secondary structure analysis by CD measurements showed 31% a-helix. The hemagglutinating activity of L. acutangula agglutinin was not inhibited by any of the monosaccharides tested. Among the disaccharides only di-N-acetylchitobiose was inhibitory. The inhibitory potency of chito-oligosaccharides increased dramatically with their size up to penta-N-acetylchltopentaose. The lectin has two binding sites for saccharides. The affinity of chito-oligosaccharides for L. acutangula lectin, as monitored by titrating the changes in the near UV-CD spectra and intrinsic fluorescence, increased strikingly with the number of GlcNAc units in them. The values of AG, AH, and A S €or the binding process showed a pronounced dependence on the size of the chito-oligosaccharides, indicating that the binding of higher oligomers is progressively more favored thermodynamically than di-N-acetylchitobiose. The thermodynamic data are consistent with an extended binding site in this lectin, which accommodates a tetrasaccharide.
Glycobiology, 2009
A simple and inexpensive method was developed to rapidly define the specificity of mannose-specific lectins toward oligomannoside-type structures. The method involved the interaction of a mixture of N-[ 14 C]-acetylated glycoasparagines, prepared by exhaustive pronase digestion of bovine pancreatic ribonuclease B and N-[ 14 C]-acetylation with [ 14 C]-acetic anhydride and containing all the possible oligomannoside-type N-glycans, with the lectin immobilized on Sepharose-4B. After exhaustive desalting, the obtained fractions were separated by high-performance thin-layer chromatography on silica gel plates and visualized by autoradiography with intensifying screen. As an example of the usefulness of this method, the fine specificity of artocarpin, the mannose-specificity lectin isolated from seeds of jackfruit (Artocarpus integrifolia) toward oligomannosidetype structures is presented. On the basis of such a determination, the best oligomannosidic ligand recognized by a mannose-specific lectin can be selected for studies of crystal structures of the lectin in complex with the defined ligand. Furthermore, some of these immobilized lectins, after definition of their precise specificities with the method, could represent valuable tools for the fractionation and characterization of oligomannose-type structures, present in complex mixtures.
Archives of Biochemistry and Biophysics, 2004
Six leguminous lectins from the seeds of plants of the Erythrina genus, namely E. caffra (ECafL), E. cristagalli (ECL), E. flabelliformis (EFL), E. lysistemon (ELysL), E. rubrinerva (ERL), and E. vespertilio (EVL), were examined to establish their sequence homology and to determine the structure and sites of attachment of their glycans. Tryptic digests of these lectins were analyzed by capillary electrophoresis coupled to electrospray mass spectrometry (CE-ESMS). Assignments were made by comparing the molecular masses of the observed tryptic peptides with those of Erythrina corallodendron lectin (ECorL), the sequence of which had been established previously. Glycan structure and genetic variations in the amino acid sequence were probed by tandem mass spectrometry. Small differences were found between the sequences of the various lectins examined and all of them exhibited Cterminal processing resulting in proteins with a C-terminal Asn residue. The major glycan of these glycoproteins was shown to be the heptasaccharide Man 3 XylFucGlcNAc 2 , consistent with previous investigations on ECL and ECorL. A minor glycan heterogeneity was observed for most lectins examined except for that of ECafL and ECorL where an extra hexose residue was observed on the reducing GlcNAc residue of the heptasaccharide.
Biochimie, 2008
Previous reports on the carbohydrate specificities of Amaranthus caudatus lectin (ACL) and peanut agglutinin (PNA, Arachis hypogea) indicated that they share the same specificity for the Thomsen-Friedenreich (T a , Galb1e3GalNAca1eSer/Thr) glycotope, but differ in monosaccharide binding e GalNAc [ Gal (inactive) for ACL; Gal [ GalNAc (weak) with respect to PNA. However, knowledge of the recognition factors of these lectins was based on studies with a small number monosaccharides and T-related oligosaccharides. In this study, a wider range of interacting factors of ACL and PNA toward known mammalian structural units, natural polyvalent glycotopes and glycans were examined by enzyme-linked lectinosorbent and inhibition assays. The results indicate that the main recognition factors of ACL, GalNAc was the only monosaccharide recognized by ACL as such, its polyvalent forms (poly GalNAca1eSer/Thr, Tn in asialo OSM) were not recognized much better. Human blood group precursor disaccharides Galb1e3/4GlcNAcb (I b /II b ) were weak ligands, while their clusters (multiantennary II b ) and polyvalent forms were active. The major recognition factors of PNAwere a combination of a or b anomers of T disaccharide and their polyvalent complexes. Although I b /II b were weak haptens, their polyvalent forms played a significant role in binding. From the 50% molar inhibition profile, the shape of the ACL combining site appears to be a cavity type and most complementary to a disaccharide of Galb1e3GalNAc (T), while the PNA binding domain is proposed to be Galb1e3GalNAca or b1as the major combining site with an adjoining subsite (partial cavity type) for a disaccharide, and most complementary to the linear tetrasaccharide, Galb1e3GalNAcb1e4Galb1e4Glc (T b 1-4L, asialo GM 1 sequence). These results should help us understand the differential contributions of polyvalent ligands, glycotopes and subtopes for the interaction with these lectins to binding, and make them useful tools to study glycosciences, glycomarkers and their biological functions. (A.M. Wu).
Structure-function relationship of monocot mannose-binding lectins
PLANT PHYSIOLOGY, 1996
The monocot mannose-binding lectins are an extended superfamily of structurally and evolutionarily related proteins, which until now have been isolated from species of the Amaryllidaceae, Alliaceae, Araceae, Orchidaceae, and Liliaceae. To explain the obvious differences in biological activities, the structure-function relationships of the monocot mannose-binding lectins were studied by a combination of glycan-binding studies and molecular modeling using the deduced amino acid sequences of the currently known lectins. Molecular modeling indicated that the number of active mannose-binding sites per monomer varies between three and zero. Since the number of binding sites is fairly well correlated with the binding activity measured by surface plasmon resonance, and is also in good agreement with the results of previous studies of the biological activities of the mannose-binding lectins, molecular modeling is of great value for predicting which lectins are best suited for a particular application. Plant lectins are an extended group of proteins that, according to a recently updated definition, comprise all plant proteins possessing at least one noncatalytic domain that binds reversibly to specific mono-or oligosaccharides (Peumans and Van Damme, 1995). Due to advances in the biochemistry and molecular biology of plant lectins during the last decade, the structural and evolutionary relationships between the different members of this apparently very , heterogeneous group of proteins have become increasingly evident. At present, the majority of all currently known plant lectins can be classified into four groups of evolutionarily related proteins: the legume lectins (Sharon and Lis, 1990), the chitin-binding lectins containing hevein domains (Raikhel and Broekaert, 1993), the type 2 ribosome-inactivating proteins (Barbieri et al., 1993), and the so-called monocot Man-binding lectins. Legume lectins are confined to species of the Leguminoseae, whereas the chitin-binding lectins and type 2 ribosome-inactivating