Structural features of the tmRNA–ribosome interaction

Abstract

Trans-translation is a process which switches the synthesis of a polypeptide chain encoded by a nonstop messenger RNA to the mRNA-like domain of a transfer-messenger RNA (tmRNA). It is used in bacterial cells for rescuing the ribosomes arrested during translation of damaged mRNA and directing this mRNA and the product polypeptide for degradation. The molecular basis of this process is not well understood. Earlier, we developed an approach that allowed isolation of tmRNA-ribosomal complexes arrested at a desired step of tmRNA passage through the ribosome. We have here exploited it to examine the tmRNA structure using chemical probing and cryo-electron microscopy tomography. Computer modeling has been used to develop a model for spatial organization of the tmRNA inside the ribosome at different stages of trans-translation.

Structural aspects of trans‐translation

“…Further steps of trans-translation cause the A79-A86 loop to occupy the mRNA-binding pocket and this position is fixed by pK1. These elements remain bound to this site during the whole trans-translation process in agreement with our chemical probing data (42). The detailed models and description for modeling process are available at: http://rnp.genebee.msu.ru/bin/ view/Projects/TmrnaEnglish.…”

“…We showed that stable secondary structure elements of the tmRNA molecule remain intact at different steps of trans-translation. The tmRNA arch becomes movable and can occupy any position between the head and the shoulder of the 30S ribosomal subunit (42). We were also able to show that one SmpB molecule remains in the tmRNA complex at different stages of trans-translation elongation (43).…”

“…We were also able to show that one SmpB molecule remains in the tmRNA complex at different stages of trans-translation elongation (43). Computer models for tmRNA organization inside the ribosome at the different stages of tmRNA passage through the ribosome were generated and verified using the chemical probing data (42). Good correlations between the experimental results and computer models allow us to extend the same approach to model the tmRNA structure in ribosomal complexes corresponding to the initial stages of trans-translation.…”

“…In our recent study, we were able to isolate tmRNA-ribosomal complexes blocked at different steps of trans-translation and to investigate the structural features of tmRNA in these complexes using chemical probing and cryo-EM tomography (42). We showed that stable secondary structure elements of the tmRNA molecule remain intact at different steps of trans-translation.…”

SmpB as the handyman of tmRNA during trans-translation

“…The model resulting from this study suggests a positioning of PK1 and H2 nearby the E-site. 76 The dynamics of the other modules are more elusive and mainly depend on when helix H5 will come out of the ribosome and unwound to be correctly translated into the mRNA path.…”