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| Title | Role of the sarcin-ricin loop of 23S rRNA in biogenesis of the 50S ribosomal subunit. |
|---|---|
| Journal, issue, pages | RNA, Vol. 31, Issue 4, Page 585-599, Year 2025 |
| Publish date | Mar 18, 2025 |
 Authors | Sepideh Fakhretaha Aval / Amal Seffouh / Kyung-Mee Moon / Leonard J Foster / Joaquin Ortega / Kurt Fredrick /   |
| PubMed Abstract | The sarcin-ricin loop (SRL) is one of the most conserved segments of ribosomal RNA (rRNA). Translational GTPases (trGTPases), such as EF-G, EF-Tu, and IF2, form contacts with the SRL that are ...The sarcin-ricin loop (SRL) is one of the most conserved segments of ribosomal RNA (rRNA). Translational GTPases (trGTPases), such as EF-G, EF-Tu, and IF2, form contacts with the SRL that are critical for GTP hydrolysis and factor function. Previous studies showed that expression of 23S rRNA lacking the SRL confers a dominant lethal phenotype in Isolated ΔSRL particles were found to be not only inactive in protein synthesis but also incompletely assembled. In particular, block 4 of the subunit, which includes the peptidyl transferase center, remained unfolded. Here, we explore the basis of this assembly defect. We find that 23S rRNA extracted from ΔSRL subunits can be efficiently reconstituted into 50S subunits, and these reconstituted ΔSRL particles exhibit full peptidyl transferase activity. We also further characterize ΔSRL particles purified from cells, using cryo-EM and proteomic methods. These particles lack density for rRNA and r-proteins of block 4, consistent with earlier chemical probing data. Incubation of these particles with excess total r-protein of the large subunit (TP50) fails to restore substantial peptidyl transferase activity. Interestingly, proteomic analysis of control and mutant particles shows an overrepresentation of multiple assembly factors in the ΔSRL case. We propose that one or more GTPases normally act to release assembly factors, and this activity is blocked in the absence of the SRL. |
 External links | RNA / PubMed:39875174 / PubMed Central |
| Methods | EM (single particle) |
| Resolution | 2.6 - 4.0 Å |
| Structure data |  EMDB-47911: 50S ribosomal subunit lacking the sarcin risin loop. Class 1.  EMDB-47913: 50S ribosomal subunit lacking the sarcin risin loop. Class 2  EMDB-47917: 50S ribosomal subunit lacking the sarcin risin loop. Class 3  EMDB-47918: 50S ribosomal subunit lacking the sarcin risin loop. Class 4  EMDB-47919: 50S ribosomal subunit lacking the sarcin risin loop. Class 5.  EMDB-47920: Control 50S ribosomal subunit for 50S subunit structures lacking the sarcin risin loop. Class 1.  EMDB-47921: Control 50S ribosomal subunit for 50S subunit structures lacking the sarcin risin loop. Class 2.  EMDB-47922: Control 50S ribosomal subunit for 50S subunit structures lacking the sarcin risin loop. Class 3. |
| Source |
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Escherichia coli (E. coli)