Conformational analysis of josamycin, a 16-membered macrolide free in solution and bound to bacterial ribosomes

The potent 14-membered macrolide antibiotics displayed a strong NMR respons e in the TRNOESY experiments whereas their metabolites which do not retain antimicrobial activity gave essentially blank TRNOESY spectra. These TRNOES Y experiments are here extended to a macrolide (josamycin) belonging to the 16-atom macrolide class in order to compare responses according to the mac rocycle size. Analysis of transferred nuclear Overhauser effect (TRNOE) exp eriments resulted in a set of constraints for all proton pairs. These const raints were used in structure determination procedures based on molecular m odelling to obtain a bound structure compatible with the experimental NMR d ata. This study allowed us to identify the binding structure at the ribosom e for the active 16-membered macrolide. The different conformations existin g in solution for this antibiotic in the free state will be compared and co rrelated with the binding conformation at the ribosome obtained with TRNOE experiments so as to establish a structure-activity relationship. The compa rative results indicate that one conformation (called S3) pre-existing in t he conformational equilibrium of macrocycle in solution and close to the ma jor one S5, is preferred in the bound state.