Engineering the Ligand Specificity of the Human Galectin-1 by Incorporation of Tryptophan Analogues

Galectin-1 is a β-galactoside-binding lectin with manifold biological functions. A single tryptophan residue (W68) in its carbohydrate binding site plays a major role in ligand binding and is highly conserved among galectins. To fine tune galectin-1 specificity, we introduced several non-canonical tryptophan analogues at this position of human galectin-1 and analyzed the resulting variants using glycan microarrays. Two variants containing 7-azatryptophan and 7-fluorotryptophan showed a reduced affinity for 3’-sulfated oligosaccharides. Their interaction with different ligands was further analyzed by fluorescence polarization competition assay. Using molecular modeling we provide structural clues that the change in affinities comes from modulated interactions and solvation patterns. Thus, we show that the introduction of subtle atomic mutations in the ligand binding site of galectin-1 is an attractive approach for fine-tuning its interactions with different ligands

Effect of Noncanonical Amino Acids on Protein–Carbohydrate Interactions: Structure, Dynamics, and Carbohydrate Affinity of a Lectin Engineered with Fluorinated Tryptophan Analogs

International audienceProtein−carbohydrate interactions play crucial roles in biology. Understanding and modifying these interactions is of major interest for fighting many diseases. We took a synthetic biology approach and incorporated noncanonical amino acids into a bacterial lectin to modulate its interactions with carbohydrates. We focused on tryptophan, which is prevalent in carbohydrate binding sites. The exchange of the tryptophan residues with analogs fluorinated at different positions resulted in three distinctly fluorinated variants of the lectin from Ralstonia solanacearum. We observed differences in stability and affinity toward fucosylated glycans and rationalized them by X-ray and modeling studies. While fluorination decreased the aromaticity of the indole ring and, therefore, the strength of carbohydrate−aromatic interactions, additional weak hydrogen bonds were formed between fluorine and the ligand hydroxyl groups. Our approach opens new possibilities to engineer carbohydrate receptor

Effect of Noncanonical Amino Acids on Protein–Carbohydrate Interactions: Structure, Dynamics, and Carbohydrate Affinity of a Lectin Engineered with Fluorinated Tryptophan Analogs

Protein–carbohydrate interactions play crucial roles in biology. Understanding and modifying these interactions is of major interest for fighting many diseases. We took a synthetic biology approach and incorporated noncanonical amino acids into a bacterial lectin to modulate its interactions with carbohydrates. We focused on tryptophan, which is prevalent in carbohydrate binding sites. The exchange of the tryptophan residues with analogs fluorinated at different positions resulted in three distinctly fluorinated variants of the lectin from <i>Ralstonia solanacearum</i>. We observed differences in stability and affinity toward fucosylated glycans and rationalized them by X-ray and modeling studies. While fluorination decreased the aromaticity of the indole ring and, therefore, the strength of carbohydrate–aromatic interactions, additional weak hydrogen bonds were formed between fluorine and the ligand hydroxyl groups. Our approach opens new possibilities to engineer carbohydrate receptors