LAUSR

Significance The cryo-electron microscopy high-resolution structures of the wild-type ribosome of the human pathogen Pseudomonas aeruginosa and its uL6 rProtein mutant, isolated from a cystic fibrosis (CF) patient, shed light on the link between a distorted initiation factor 2 (IF2) binding site, a deletion in uL6, and a 50-Å distal H69–h44 B2a&b intersubunit bridges. These cumulative structural alterations interfere with the initiation of proper protein synthesis. Here, via efforts at understanding a specific and so-far-unknown aminoglycoside (AG)-resistant mechanism, we widen the concept of diversity of resistance mechanisms. Thus, we reveal a defective ribosome obtained by a conformational change of essential ribosomal intersubunit bridges and an alteration of the secondary AG binding site. Resistance to antibiotics has become a major threat to modern medicine. The ribosome plays a fundamental role in cell vitality by the translation of the genetic code into proteins; hence, it is a major target for clinically useful antibiotics. We report here the cryo-electron microscopy structures of the ribosome of a pathogenic aminoglycoside (AG)-resistant Pseudomonas aeruginosa strain, as well as of a nonresistance strain isolated from a cystic fibrosis patient. The structural studies disclosed defective ribosome complex formation due to a conformational change of rRNA helix H69, an essential intersubunit bridge, and a secondary binding site of the AGs. In addition, a stable conformation of nucleotides A1486 and A1487, pointing into helix h44, is created compared to a non-AG-bound ribosome. We suggest that altering the conformations of ribosomal protein uL6 and rRNA helix H69, which interact with initiation-factor IF2, interferes with proper protein synthesis initiation.