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TitleInteractions between terminal ribosomal RNA helices stabilize the large ribosomal subunit.
Journal, issue, pagesRNA, Vol. 29, Issue 10, Page 1500-1508, Year 2023
Publish dateJul 7, 2023
AuthorsAmos J Nissley / Tammam S Kamal / Jamie H D Cate /
PubMed AbstractThe ribosome is a large ribonucleoprotein assembly that uses diverse and complex molecular interactions to maintain proper folding. In vivo assembled ribosomes have been isolated using MS2 tags ...The ribosome is a large ribonucleoprotein assembly that uses diverse and complex molecular interactions to maintain proper folding. In vivo assembled ribosomes have been isolated using MS2 tags installed in either the 16S or 23S ribosomal RNAs (rRNAs), to enable studies of ribosome structure and function in vitro. RNA tags in the 50S subunit have commonly been inserted into an extended helix H98 in 23S rRNA, as this addition does not affect cellular growth or in vitro ribosome activity. Here, we find that 50S subunits with MS2 tags inserted in H98 are destabilized compared to wild-type (WT) 50S subunits. We identify the loss of RNA-RNA tertiary contacts that bridge helices H1, H94, and H98 as the cause of destabilization. Using cryogenic electron microscopy (cryo-EM), we show that this interaction is disrupted by the addition of the MS2 tag and can be restored through the insertion of a single adenosine in the extended H98 helix. This work establishes ways to improve MS2 tags in the 50S subunit that maintain ribosome stability and investigates a complex RNA tertiary structure that may be important for stability in various bacterial ribosomes.
External linksRNA / PubMed:37419664 / PubMed Central
MethodsEM (single particle)
Resolution1.84 - 1.98 Å
Structure data

29449

8fto

EMDB-29449: E. coli 70S ribosome with an improved MS2 Tag inserted in H98 (Composite Map)
PDB-8fto: E. coli 70S ribosome with an improved MS2 tag inserted in H98
Method: EM (single particle) / Resolution: 1.85 Å

EMDB-29483: E. coli 70S ribosome with an improved MS2 tag inserted in H98 (70S map)
Method: EM (single particle) / Resolution: 1.85 Å

EMDB-29484: E. coli 70S ribosome with an improved MS2 tag inserted in H98 (50S focused map)
Method: EM (single particle) / Resolution: 1.84 Å

EMDB-29485: E. coli 70S ribosome with an improved MS2 tag inserted in H98 (30S focused map)
Method: EM (single particle) / Resolution: 1.98 Å

Chemicals

ChemComp-MG:
Unknown entry

ChemComp-FME:
N-FORMYLMETHIONINE / N-Formylmethionine

ChemComp-HOH:
WATER / Water

Source
  • escherichia coli (E. coli)
KeywordsRIBOSOME / MS2-tag / H98

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