Structures of Mycobacterium smegmatis 70S ribosomes in complex with HPF, tmRNA, and P-tRNA

Satabdi Mishra,Anu Tyagi,Tofayel Ahmed,Jian Shi,Shashi Bhushan

doi:10.1038/s41598-018-31850-3

Satabdi Mishra, Anu Tyagi + Show 3 more

Open Access

https://doi.org/10.1038/s41598-018-31850-3

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Abstract

Ribosomes are the dynamic protein synthesis machineries of the cell. They may exist in different functional states in the cell. Therefore, it is essential to have structural information on these different functional states of ribosomes to understand their mechanism of action. Here, we present single particle cryo-EM reconstructions of the Mycobacterium smegmatis 70S ribosomes in the hibernating state (with HPF), trans-translating state (with tmRNA), and the P/P state (with P-tRNA) resolved to 4.1, 12.5, and 3.4 Å, respectively. A comparison of the P/P state with the hibernating state provides possible functional insights about the Mycobacteria-specific helix H54a rRNA segment. Interestingly, densities for all the four OB domains of bS1 protein is visible in the hibernating 70S ribosome displaying the molecular details of bS1-70S interactions. Our structural data shows a Mycobacteria-specific H54a-bS1 interaction which seems to prevent subunit dissociation and degradation during hibernation without the formation of 100S dimer. This indicates a new role of bS1 protein in 70S protection during hibernation in Mycobacteria in addition to its conserved function during translation initiation.

Highlights

Ribosomes are the protein synthesis machineries of the cell, which translate genetic codes on mRNA into protein in all living species
Ribosomes were prepared from M. smegmatis (Ms) cells harvested at the log (70S-log) and stationary phases (70S-stat) to obtain the 70S ribosomes in different functional states
The 70S-log Cryo-electron microscopy (cryo-EM) dataset resulted in one major class of 70S ribosome resolved to 3.4 Å and one minor class of trans-translating 70S ribosome with the presence of a transfer-messenger RNA (tmRNA)-small protein B (SmpB) (0.6% particles) resolved to 12.5 Å (Supplementary Figs S1–S3)

Summary

Introduction

Ribosomes are the protein synthesis machineries of the cell, which translate genetic codes on mRNA into protein in all living species. We observe a novel interaction of Ms bS1 protein with Mycobacteria-specific helix H54a in the hibernating state indicating an additional function of bS1 in stabilization and protection of the Ms 70S ribosome during stationary/dormant phase. The 70S-log cryo-EM dataset resulted in one major class of 70S ribosome (with P-tRNA, 99.4% particles) resolved to 3.4 Å (the P/P state) and one minor class of trans-translating 70S ribosome with the presence of a tmRNA-SmpB (0.6% particles) resolved to 12.5 Å (Supplementary Figs S1–S3).

Results

Conclusion