A glycosaminoglycan-binding LecA-like lectin from Xenorhabdus nematophila: structural and biophysical characterization.

Glycosaminoglycan (GAG)-binding lectins represent a rare class of carbohydrate-binding proteins with the ability to recognize and organize linear polysaccharide chains. Here, we describe XN-IL, a novel calcium-dependent lectin from the Gram-negative bacterium Xenorhabdus nematophila, which exhibits an unusual specificity for glycosaminoglycans. X. nematophila is an entomopathogenic bacterium and a symbiont of insect-parasitic Steinernema nematodes. Glycan array screening, analytical ultracentrifugation, and differential scanning fluorimetry revealed that XN-IL selectively binds hyaluronan and low-sulfated heparan sulfate, while showing negligible affinity for monosaccharides and galactosylated glycans. GAG binding is mediated exclusively by calcium ions, enabling the reversible crosslinking and precipitation of hyaluronan polymers. Crystal structures of the apo and ligand-bound forms reveal a conserved LecA-like fold with a widened, calcium-dependent binding pocket that accommodates extended GAG chains without major conformational rearrangements. XN-IL is the first member of the LecA family with defined GAG specificity and the first lectin identified in the genus Xenorhabdus. Its divergence from galactophilic LecA homologues reflects an evolutionary adaptation towards calcium-driven recognition and reversible assembly of linear polysaccharides. These findings expand the functional diversity of the LecA family and introduce XN-IL as a new tool for probing and manipulating GAG-based polymer systems.