The delineation of the physiological significance of protein (lectin)-glycan recognition and the structural analysis of individual lectins have directed our attention to studying them in combination. In this report, we tested the hypothesis of hybrid formation by using binary mixtures of homodimeric galectin-1 and -7 as well as a proteolytically truncated version of chimera-type galectin-3. Initial supportive evidence is provided by affinity chromatography using resin-presented galectin-7. Intriguingly, the extent of cell binding by cross-linking of surface counter-receptor increased significantly for monomeric galectin-3 form by the presence of galectin-1 or -7. Pulsed-field gradient NMR (nuclear magnetic resonance) diffusion measurements on these galectin mixtures indicated formation of heterodimers as opposed to larger oligomers.15N-1H heteronuclear single quantum coherence NMR spectroscopy and molecular dynamics simulations allowed us to delineate how different galectins interact in the heterodimer. The possibility of domain exchange between galectins introduces a new concept for understanding the spectrum of their functionality, particularly when these effector molecules are spatially and temporally co-expressed as found in vivo.
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Even though our NMR data indicate that Gal-7 interacts with Gal-1, effects appear to be relatively weak. We explain this by considering that homodimers of Gal-1 (unlike Gal-7) are very stable, such that its monomer population is naturally low, making NMR detection of Gal-1-involved hetero-oligomers more difficult due to its lower population. This also suggests that the Gal-1 : Gal-7 heterodimer Kd is larger than the Gal-1 homodi-mer Kd, which is in any event less that 1 μM. This view is supported by variant Gal-1 [8S] and tandem-repeat-type Gal-8. When the Gal-1 CRDs are not strongly associated, as in the Gal-1 [8S] variant , hybrid interactions with Gal-7 or the truncated Gal-3 CRD occur more readily than with wild-type Gal-1, as shown in our gel electrophoresis results (Figure 2). Furthermore, tandem-repeat-type Gal-8, with its two monomeric CRDs, also readily associates with resin-bound Gal-7 (Figure 2F), unlike wild-type Gal-1.
NMR experiments were performed at the Minnesota NMR Center with funding for NMR instrumentation provided by the Office of the Vice-President for Research, the University of Minnesota Medical School and College of Biological Sciences, as well as generous funding from the US National Institutes of Health, the US National Science Foundation, and the Minnesota Medical Foundation. Insightful discussions with Drs. B. Friday and A. Leddoz, as well as valuable recommendations from the reviewers, are gratefully acknowledged.
© 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.