Selective Protein Synthesis by Ribosomes with a Drug-Obstructed Exit Tunnel

Abstract

The polypeptide exit tunnel is an important functional compartment of the ribosome where the newly synthesized proteins are surveyed. The tunnel is the target of clinically important macrolide antibiotics. Macrolides plug the tunnel and are believed to stop production of all proteins. Contrary to this view, we show that drug-bound ribosomes can synthesize a distinct subset of cellular polypeptides. The structure of a protein defines its ability to thread through the antibiotic-obstructed tunnel. Synthesis of certain polypeptides that initially bypass translational arrest can be stopped at later stages of elongation while translation of some proteins goes to completion. Our findings reveal that small-molecule effectors can accentuate the discriminatory properties of the ribosomal exit tunnel and that macrolide antibiotics reshape the cellular proteome rather than block global protein synthesis.

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

“…Our finding that CHL and LZD act at the defined locations within the gene extends the concept of context-specific action of ribosome-targeting inhibitors (32). Macrolides, which bind to the nascent peptide exit tunnel, arrest translation at a limited number of codons within the ORF also depending on the nature of the nascent peptide and of the incoming amino acid (27,28,32,38).…”

“…As a result, PTC inhibitors halt translation at multiple locations along the gene, whereas macrolides arrest ribosome only at a limited number of codons within the ORF, and synthesis of the proteins lacking the "problematic" sequences may not be inhibited at all (32,39). For this reason, cells treated with excess of macrolide antibiotics continue to synthesize a limited subset of polypeptides (32), whereas protein translation in cells exposed to CHL or LZD is essentially abolished (Fig.…”

“…Macrolides, which bind to the nascent peptide exit tunnel, arrest translation at a limited number of codons within the ORF also depending on the nature of the nascent peptide and of the incoming amino acid (27,28,32,38). The context requirements for macrolides, however, are principally different from those for the PTC-targeting antibiotics.…”

“…To further examine the influence of the sequence context on druginduced arrest, we selected a well-defined model system appealingly suited for cell-free translation experiments (32). According to the ribosome-profiling data, the drug-induced ribosome stalling at the Leu5 codon of the hns gene represents one of the 10 strongest arrest sites common for both CHL and LZD (Fig.…”

Targeting Antibiotic Resistance

“…In fact, modeling studies have shown that even with a bound macrolide, the tunnel can still accommodate a nascent peptide chain. The macrolide nevertheless greatly hinders the progression of the peptide, which is usually dissociated by the peptidyl‐tRNA drop‐off mechanism before reaching its full size 51, 52. Macrolides have also been shown to block the formation of the large 50S ribosomal subunit by binding to its precursors 53…”