Structures of the Escherichia coli ribosome with antibiotics bound near the peptidyl transferase center explain spectra of drug action

Abstract

Differences between the structures of bacterial, archaeal, and eukaryotic ribosomes account for the selective action of antibiotics. Even minor variations in the structure of ribosomes of different bacterial species may lead to idiosyncratic, species-specific interactions of the drugs with their targets. Although crystallographic structures of antibiotics bound to the peptidyl transferase center or the exit tunnel of archaeal ( Haloarcula marismortui ) and bacterial ( Deinococcus radiodurans ) large ribosomal subunits have been reported, it remains unclear whether the interactions of antibiotics with these ribosomes accurately reflect those with the ribosomes of pathogenic bacteria. Here we report X-ray crystal structures of the Escherichia coli ribosome in complexes with clinically important antibiotics of four major classes, including the macrolide erythromycin, the ketolide telithromycin, the lincosamide clindamycin, and a phenicol, chloramphenicol, at resolutions of ∼3.3 Å –3.4 Å . Binding modes of three of these antibiotics show important variations compared to the previously determined structures. Biochemical and structural evidence also indicates that interactions of telithromycin with the E. coli ribosome more closely resembles drug binding to ribosomes of bacterial pathogens. The present data further argue that the identity of nucleotides 752, 2609, and 2055 of 23S ribosomal RNA explain in part the spectrum and selectivity of antibiotic action.

Recruitment of a species-specific translational arrest module to monitor different cellular processes

“…When mifM-myc was translated with Bs hybrid PURE system, the 30 kDa band again appeared earlier (at 5 min; lane 12) than the other major band, which now migrated at the position of 12 kDa and became detectable at 10 min (lanes [13][14][15]. This 12 kDa band must represent the full-length MifM-myc polypeptide because it is resistant to RNaseA (lanes [18][19][20] and uniquely observed after the myc-tag addition (compare lanes 2-5 and lanes 12-15). RNaseA treatment of the 5-min sample led to the disappearance of the 30 kDa band with concomitant appearance of the 10 kDa band (lane 17), indicating that the former (MifM′-tRNA) was converted to the latter (polypeptide moiety, MifM′).…”

“…Because at least approximately 90% of the translation products of MifM-myc were in the form of MifM′-tRNA 5 min after the start of the reaction, we conclude that most, if not all, of the ribosomes that translate MifM-myc had undergone translational pause. However, the arrest seems to be released spontaneously at a certain frequency, leading to the production of the full-length MifM-myc protein at the later time points of the incubation (lanes [18][19][20]. The ratio of arrest/full-length went down markedly during the early part of the time course, suggesting that the arrest species was not a deadend product but an intermediate in the synthesis of the full-length protein.…”

“…An important unanswered question is the species specificity of the arrest sequences, whose known examples are quite divergent in amino acid sequences. Because the ribosome is thought to be one of the most universal enzymes, it is conceivable that different arrest sequences still interact with conserved sets of ribosomal components, as observed with a number of antibiotics (19,20). Alternatively, they might interact with the ribosome in disparate ways, at least in the initial recognition.…”

Targeting Antibiotic Resistance

“…Therefore, questions remained as to the actual binding mode of solithromycin and telithromycin in pathogenic bacteria. In 2010, Cate, Mankin, and co‐workers crystallized solithromycin as well as telithromycin in complex with the E. coli ribosome and were able to indicate that the placement of these ketolides most likely reflects the binding in S. aureus ribosomes given the sequence conservation of the A752:U2609 base pair among many eubacteria 71, 72…”

Context-specific inhibition of translation by ribosomal antibiotics targeting the peptidyl transferase center

“…CHL binds to the PTC A site, preventing proper placement of the aminoacyl moiety of aa-tRNA ( Fig. 1) (10)(11)(12). Because the key interactions with the ribosome in the A site are shared by all aa-tRNAs, CHL is viewed as a universal inhibitor of peptide bond formation (reviewed in ref.…”