A lectin from Platypodium elegans with unusual specificity and affinity for asymmetric complex N-glycans.
Lectin activity with specificity for mannose and glucose has been detected in the seed of Platypodium elegans, a legume plant from the Dalbergieae tribe. The gene of Platypodium elegans lectin A has been cloned, and the resulting 261-amino acid protein belongs to the legume lectin family with similarity with Pterocarpus angolensis agglutinin from the same tribe. The recombinant lectin has been expressed in Escherichia coli and refolded from inclusion bodies. Analysis of specificity by glycan array evidenced a very unusual preference for complex type N-glycans with asymmetrical branches. A short branch consisting of one mannose residue is preferred on the 6-arm of the N-glycan, whereas extensions by GlcNAc, Gal, and NeuAc are favorable on the 3-arm. Affinities have been obtained by microcalorimetry using symmetrical and asymmetrical Asn-linked heptasaccharides prepared by the semi-synthetic method. Strong affinity with K(d) of 4.5 μm was obtained for both ligands. Crystal structures of Platypodium elegans lectin A complexed with branched trimannose and symmetrical complex-type Asn-linked heptasaccharide have been solved at 2.1 and 1.65 Å resolution, respectively. The lectin adopts the canonical dimeric organization of legume lectins. The trimannose bridges the binding sites of two neighboring dimers, resulting in the formation of infinite chains in the crystal. The Asn-linked heptasaccharide binds with the 6-arm in the primary binding site with extensive additional contacts on both arms. The GlcNAc on the 6-arm is bound in a constrained conformation that may rationalize the higher affinity observed on the glycan array for N-glycans with only a mannose on the 6-arm.
Centre de Recherche sur les Macromolécules Végétales-CNRS (affiliated with Université Joseph Fourier and Institut de Chimie Moléculaire de Grenoble), 38041 Grenoble, France.
Raquel Guimarães
RG
Benevides
Géraldine
G
Ganne
Rafael da Conceição
Rda C
Simões
Volker
V
Schubert
Mathäus
M
Niemietz
Carlo
C
Unverzagt
Valérie
V
Chazalet
Christelle
C
Breton
Annabelle
A
Varrot
Benildo Sousa
BS
Cavada
Anne
A
Imberty
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002151:Calorimetry; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D007887:Fabaceae / Q000737:chemistry*; D006384:Hemagglutination / Q000187:drug effects; D006801:Humans; D006860:Hydrogen Bonding; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010936:Plant Extracts / Q000737:chemistry* / Q000494:pharmacology; D037121:Plant Lectins / Q000737:chemistry* / Q000494:pharmacology; D011134:Polysaccharides / Q000138:chemical synthesis / Q000737:chemistry*; D040081:Protein Array Analysis; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012639:Seeds / Q000737:chemistry*; D013816:Thermodynamics
D010936:Plant Extracts; D037121:Plant Lectins; D011134:Polysaccharides