CONFORMATION OF DEOXY OLIGOSACCHARIDES - SOLUTION CONFORMATION AND FLEXIBILITY OF THE ANTIBIOTIC KIJANIMICIN STUDIED BY NMR-SPECTROSCOPY AND MMC CALCULATIONS

The solution conformation of the macrolide antibiotic kijanimicin (1) was deduced by a combination of NMR spectroscopy and potential energy calculations of the carbohydrate parts using the HSEA force field. The study was based on temperature dependent NMR spectroscopic shift data , 1D nuclear Overhauser enhancements (NOE) of selected protons and 2D rotating frame NOE of the complete antibiotic. NOEs in the oligosaccha ride part of the antibiotic indicated some unusual short distances bet ween the alpha-linked digitoxose residues A and B, as well as between not directly linked residues A and C. Calculation of preferred conform ations revealed a global minimum with PHI(A,B)/PSI(A,B) almost-equal-t o 70-degrees/50-degrees, PHI(B,C)/PSI(B,C) almost-equal-to 60-degrees/ 30-degrees, and PHI(B,D)/PSI(B,D) almost-equal-to -60-degrees/5-degree s for the oligosaccharide moiety. MMC simulations of the conformationa l flexibility indicated a high rigidity of the carbohydrate. The range of the glycosidic angles covered during MMC simulations was 50-degree s-70-degrees for the alpha-linkages of the oligosaccharide. The beta-g lycosidic linkages of residue D and E showed slightly more flexibility , covering a range of 120-degrees-270-degrees. Minimum energy calculat ions of the oligosaccharide part revealed a second minimum with differ ent orientation of sugar residues A-B and an energy difference of 2 kc al/mol to the global minimum. This energy difference increased to 12 k cal/mol when the complete antibiotic was calculated and MMC simulation s did not show any transitions between the two minima. The experimenta l NOE in the oligosaccharide moiety and the enhancements between carbo hydrate and lactone part agree well with theoretical NOEs that were ob tained by sampling the global minimum in MMC simulations. The conforma tion of the macrolide moiety was evaluated by comparing experimental N OEs with the crystal structure of the isolated aglycon. The conformati on found in solution is slightly altered around the sites of attachmen t of the carbohydrate parts. Constancy of the proton chemical shifts o ver a temperature range of 70-degrees-C indicated the high degree of r igidity of the carbohydrate moiety and most of the macrolide structure . Flexibility was observed for the C-13/C-20 part of the lactone ring. The antibiotic has a fairly unpolar surface and adopts a well defined and quite rigid conformation.