Synthesis of fused triazoles as probes for the active site of retaining beta-glycosidases: From which direction is the glycoside protonated?

The structure of the beta-glycosidase inhibitors 1-7 and 10-13 suggests that protonation of O-C(1) (the glycosidic O-center) of the substrate by a carboxy group of the retaining beta-glycosidases does not occur in the plane perpendicular to the ring of the glycon (beta-side; 'from the top'), but in the plane of the ring ('from the side'). The triazoles 17 and 18 have been prepared in six steps from the L-xylofuranose 21. They possess a CH group instead of the N-center of the related tetrazoles 4 and 5, corresponding to the glycosidic O-atom, and a very similar structure, both in solution and in the solid state. Unlike the tetrazoles, however, which are good-to-medium inhibitors of retaining beta-glycosidases, the triazoles do not inhibit the beta-glycosidases from sweet almonds, snail, and bovine liver, and only slightly inhibit the beta-glucosidase From Caldocellum saccharolyticum. This is in keeping with the proposed direction of protonation in the plane of the saccharide ring and with modelling studies, docking 4 into the active site of the white clover cyanogenic beta-glucosidase and 6 into the E. coli beta-galactosidase and the Lactococcus lactis 6-phospho-beta-galactosidase.