α-galactomannan Davanat binds galectin-1 at a site different from the conventional galectin carbohydrate binding domain

• Published in: Glycobiology (Oxford) 2009-09, Vol.19 (9), p.1034-1045
• Main Authors: Miller, Michelle C, Klyosov, Anatole, Mayo, Kevin H
• Format: Article
• Language: English
• Subjects: Binding Sites; Carbohydrate Sequence; diffusion; Galectin 1 - metabolism; glycan; lectin; Magnetic Resonance Spectroscopy; Mannans - metabolism; Models, Molecular; Molecular Sequence Data; NMR spectroscopy; Original; protein Q
• ISSN: 0959-6658; 1460-2423
• DOI: 10.1093/glycob/cwp084

Galectins are a sub-family of lectins, defined by their highly conserved β-sandwich structures and ability to bind to β-galactosides, like Gal β1-4 Glc (lactose). Here, we used ¹⁵N-¹H HSQC and pulse field gradient (PFG) NMR spectroscopy to demonstrate that galectin-1 (gal-1) binds to the relatively large galactomannan Davanat, whose backbone is composed of β1-4-linked D-mannopyranosyl units to which single D-galactopyranosyl residues are periodically attached via α1-6 linkage (weight-average MW of 59 kDa). The Davanat binding domain covers a relatively large area on the surface of gal-1 that runs across the dimer interface primarily on that side of the protein opposite to the lactose binding site. Our data show that gal-1 binds Davanat with an apparent equilibrium dissociation constant (Kd) of 10 x 10⁻⁶ M, compared to 260 x 10⁻⁶ M for lactose, and a stiochiometry of about 3 to 6 gal-1 molecules per Davanat molecule. Mannan also interacts at the same galactomannan binding domain on gal-1, but with at least 10-fold lower avidity, supporting the role of galactose units in Davanat for relatively strong binding to gal-1. We also found that the β-galactoside binding domain remains accessible in the gal-1/Davanat complex, as lactose can still bind with no apparent loss in affinity. In addition, gal-1 binding to Davanat also modifies the supermolecular structure of the galactomannan and appears to reduce its hydrodynamic radius and disrupt inter-glycan interactions thereby reducing glycan-mediated solution viscosity. Overall, our findings contribute to understanding gal-1-carbohydrate interactions and provide insight into gal-1 function with potentially significant biological consequences.