The crystal structure of Nictaba reveals its carbohydrate-binding properties and a new lectin dimerization mode

Nictaba is a (GlcNAc)n-binding, stress-inducible lectin from Nicotiana tabacum that serves as a representative for the family of Nictaba-related lectins, a group of proteins that play pivotal roles in plant defense mechanisms and stress response pathways. Despite extensive research into the biological activities and physiological role(s) of the lectin, the three-dimensional structure of Nictaba remained largely unknown. Here, we report crystal structures for Nictaba in the apo form and bound to chitotriose. The structures reveal a jelly-roll fold for the Nictaba protomer similar to the cucumber Cus17 lectin, but show an unprecedented dimerization mode among all structurally characterized lectins to date. The chitotriose binding mode, similar to Cus17, centers around the central GlcNAc residue, providing insights into the determinants of specificity of Nictaba towards carbohydrate structures. By integrating these structural insights with inputs from glycan arrays, molecular docking, and molecular dynamics simulations, we propose that Nictaba employs a single carbohydrate-recognition domain within each of the two subunits in the dimer to display pronounced specificity towards GlcNAc-containing carbohydrates. Furthermore, we identified amino acid residues involved in the extended binding site capable of accommodating structurally diverse high-mannose and complex N-glycans. Glycan array and in silico analyses revealed interactions centered around the conserved Man3GlcNAc2 core, explaining the broad recognition of N-glycan structures. Collectively, the structural and biochemical insights presented here fill a hitherto substantial void into the atlas of lectin structure-function relationships and pave the way for future developments in plant stress biology and lectin-based applications.