RESTRICTED FLEXIBILITY OF THE GLYCOSIDIC LINKAGE IN ALPHA(1-]3)-L-DIGITOXOSIDES AS DETERMINED BY NMR-SPECTROSCOPY AND MMC CALCULATIONS

The preferred conformations of nine protected alpha(1-->3)-linked digi toxoside trisaccharides 1-9, L-Dig-(1-3)-alpha-L-Dig-(1-3)-alpha-L-Dig -(C-B-A), constituents of tetronolide antibiotics, were determined by a combination of 300-MHz NMR spectroscopy and potential energy calcula tions using the GE-GOP programme. Chemical shift differences and nucle ar Overhauser enhancements indicated some unusual short distances betw een not directly bound digitoxose residues A and C. Thus, the glycosid ic linkages of the nine alpha(1-->3)-linked digitoxosides have a fairl y high rigidity as indicated by the relatively large absolute NOE (app roximate to 3%) from 1C-H to 5A-H. MMC simulations of the conformation al flexibility resulted in preferred conformations of the glycosidic L inkages of Phi(C,B)/Psi Psi(C,B) approximate to 70 degrees/40 degrees and Phi(B,A/)Psi(B,A) approximate to 65 degrees/50 degrees with ranges of 40 degrees/50 degrees and 50 degrees/60 degrees. By energy minimis ations a second minimum was identified for the B-A glycosidic linkage at Phi(B,A)Psi(B,A) approximate to 0 degrees/40 degrees with an energy higher than 2.0 kcal/mol to the main minimum. The MMC-derived ensembl e-averaged NOEs that were obtained by sampling the main minimum agree well with the experimental values, whereas NOEs derived from MMC runs that stayed at the local minima are not in accord with experimental va lues. The MMC simulations failed to give transitions between the two m inima presumably because of a high energy barrier between the minima. All nine trisaccharides seem to adopt a well-defined conformation char acterised by a narrow distribution in the conformational space that is located in the region of the global minimum.