Dataset Information

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

ABSTRACT: Galectins are a sub-family of lectins, defined by their highly conserved beta-sandwich structures and ability to bind to beta-galactosides, like Gal beta1-4 Glc (lactose). Here, we used (15)N-(1)H 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 beta1-4-linked d-mannopyranosyl units to which single d-galactopyranosyl residues are periodically attached via alpha1-6 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 (K(d)) 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 beta-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.

SUBMITTER: Miller MC

PROVIDER: S-EPMC2720280 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

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

Miller Michelle C MC Klyosov Anatole A Mayo Kevin H KH

Glycobiology 20090618 9

Galectins are a sub-family of lectins, defined by their highly conserved beta-sandwich structures and ability to bind to beta-galactosides, like Gal beta1-4 Glc (lactose). Here, we used (15)N-(1)H 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 beta1-4-linked d-mannopyranosyl units to which single d-galactopyranosyl residues are periodically attached via alpha1-6 linkage ...[more]

PMID: 19541770

Similar Datasets

Project description:gal-1 (galectin-1) mediates cell-cell and cell-extracellular matrix adhesion, essentially by interacting with beta-galactoside-containing glycans of cell-surface glycoconjugates. Although most structural studies with gal-1 have investigated its binding to simple carbohydrates, in particular lactose and N-acetyl-lactosamine, this view is limited, because gal-1 functions at the cell surface by interacting with more complex glycans that are heterogeneous in size and composition. In the present study we used NMR spectroscopy to investigate the interaction of human gal-1 with a large (120 kDa) complex glycan, GRG (galactorhamnogalacturonate glycan), that contains non-randomly distributed mostly terminal beta(1-->4)-linked galactose side chains. We used 15N-1H-HSQC (heteronuclear single quantum coherence) NMR experiments with 15N-enriched gal-1 to identify the GRG-binding region on gal-1 and found that this region covers a large surface area on gal-1 that includes the quintessential lactose-binding site and runs from that site through a broad valley or cleft towards the dimer interface. HSQC and pulsed-field-gradient NMR diffusion experiments also show that gal-1 binds GRG with a gal-1:GRG stoichiometry of about 5:1 (or 6:1) and with average macroscopic and microscopic equilibrium dissociation constants (Kd) of 8 x 10(-6) M and 40 x 10(-6) M (or 48 x 10(-6) M) respectively, indicating stronger binding than to lactose (Kd=520 x 10(-6) M). Although gal-1 may bind GRG in various ways, the glycan can be competed for by lactose, suggesting that there is one major mode of interaction. Furthermore, even though terminal motifs on GRG are Gal-beta(1-->4)-Gal rather than the traditional Gal-beta(1-->4)-Glc/GlcNAc (where GlcNAc is N-acetylglucosamine), we show that the disaccharide Gal-beta(1-->4)-Gal can bind gal-1 at the lactose-binding domain. In addition, gal-1 binding to GRG disrupts inter-glycan interactions and decreases glycan-mediated solution viscosity, a glycan decongestion effect that may help explain why gal-1 promotes membrane fluidity and lateral diffusion of glycoconjugates within cell membranes. Overall, our results provide an insight into the function of galectin in situ and have potential significant biological consequences.

Dataset Information

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

Publications

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

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets