Perdeuteration, crystallization, data collection and comparison of five neutron diffraction data sets of complexes of human galectin-3C.

Galectin-3 is an important protein in molecular signalling events involving carbohydrate recognition, and an understanding of the hydrogen-bonding patterns in the carbohydrate-binding site of its C-terminal domain (galectin-3C) is important for the development of new potent inhibitors. The authors are studying these patterns using neutron crystallography. Here, the production of perdeuterated human galectin-3C and successive improvement in crystal size by the development of a crystal-growth protocol involving feeding of the crystallization drops are described. The larger crystals resulted in improved data quality and reduced data-collection times. Furthermore, protocols for complete removal of the lactose that is necessary for the production of large crystals of apo galectin-3C suitable for neutron diffraction are described. Five data sets have been collected at three different neutron sources from galectin-3C crystals of various volumes. It was possible to merge two of these to generate an almost complete neutron data set for the galectin-3C-lactose complex. These data sets provide insights into the crystal volumes and data-collection times necessary for the same system at sources with different technologies and data-collection strategies, and these insights are applicable to other systems.

Arene-anion based arginine-binding motif on a galactose scaffold: structure-activity relationships of interactions with arginine-rich galectins.

Two series of C3-benzamido and O2-anion-substituted galactopyranosides were synthesized and studied as binders to arginine-rich proteins galectin-1, -3, -7, -8N (N-terminal domain), and -9N (N-terminal domain). The first series had a 4-methylbenzamide at C3 and the anionic O2-substituent was varied. The second series varied the 4-substituent of the C3-benzamide, whereas the anionic O2 substituent was kept as a sulfate. The influence of the O2-anion substituent correlated negatively with the oxygen charge density in case of galectin-1, -3, and -9N. In the second series, the electron-donating capacity of the 4-substituent of the C3-benzamides correlated positively with the magnitude of the affinity enhancement by the 2O-sulfate.

Synthesis of 3-azido-3-deoxy-beta-D-galactopyranosides.

Three efficient routes to 3-azido-3-deoxy-beta-D-galactopyranosides were developed relying on a double inversion protocol at C3. Two of the routes were demonstrated to work with both O- and S-glycosides. In all three routes, the 2-O-acetyl-3-azido-4,6-O-benzylidene-3-deoxy-beta-D-galactopyranosides were obtained by an azide inversion of the key intermediates 2-O-acetyl-4,6-O-benzylidene-3-O-trifluoromethanesulfonyl-beta-D-gulopyranosides. The intermediate gulopyranosides were in turn obtained from 2-O-acetyl-4,6-O-benzylidene-3-O-trifluoromethanesulfonyl-beta-D-galactopyranosides, installed in one pot from the 4,6-O-benzylidene-beta-D-galactopyranosides, by inversion with nitrite or acetate. For O-glycosides, the gulopyranoside configuration could alternatively be obtained from the 4,6-O-benzylidene-beta-D-galactopyranoside by elimination to give the 2,3-dianhydro derivative followed by a highly stereoselective cis-dihydroxylation.