Good Vibrations: Structural Remodeling of Maturing Yeast Pre-40S Ribosomal Particles Followed by Cryo-Electron Microscopy
Abstract
:1. Introduction
2. Results
2.1. Proteomic Characterization of Tsr1-purified pre-40S Particles
2.2. Cryo-EM Analysis of Tsr1-FPZ pre-40S Particles
2.3. Structural Heterogeneity of the Head Domain
2.4. A “Wriggling” of the Platform Domain
3. Discussion
3.1. Cryo-EM Structures Suggest that pre-40S Particles Transit through a Vibrating State
3.2. Dim1 Activity and Release from pre-40S Particles Might Correspond to Early Cytoplasmic Events
3.3. Ltv1 Conformational Change and Beak Wriggling Precede Ltv1 Phosphorylation by Hrr25
4. Material and Methods
4.1. Purification of Cytoplasmic pre-40S Particles
4.2. Protein Digestion and NanoLC-MS/MS Analysis
4.3. Bioinformatic MS Data Analysis
4.4. Grid Preparation and cryo-EM Images Acquisition
4.5. Single Particle Analysis
4.6. Cryo-EM Maps Interpretation
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
Data Availability
References
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Sample Availability: Yeast strains harboring a tagged version of Tsr1 are available from the authors for pre-40S particles purification. |
Protein | Peptide Sequence | Phosphorylation Protein Position | % Phosphosite run 1 | % Phosphosite run 2 | % Phosphosite Average | |
---|---|---|---|---|---|---|
Tsr1 | DTLDEYAPEGTEDWSDYDEDFEYDGLTTAR | T354 or S358 | 100 | 100 | 0 | Non phosphorylated |
100 | Phosphorylated | |||||
Ltv1 | GAMSDVSGFSMSSSAIAR | S336 or S339 | 7.8 | 5.2 | 93.5 | Non phosphorylated |
6.5 | Phosphorylated | |||||
Ltv1 | VTNTLSSLKF | S460 | 0 | 2.5 | 98.7 | Non phosphorylated |
1.3 | Phosphorylated | |||||
Enp1 | EKESQVEDMQDDEPLANEQNTSR | S172 | 0 | 4.3 | 97.9 | Non phosphorylated |
2.1 | Phosphorylated | |||||
Enp1 | ESQVEDMQDDEPLANEQNTSRGNISSGLK | T189 or S195 or S196 | 0 | 1.2 | 99.4 | Non phosphorylated |
0.6 | Phosphorylated | |||||
Enp1 | ILDDGSNGEDATR | S404 | 0.2 | 0.2 | 99.8 | Non phosphorylated |
0.2 | Phosphorylated |
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Shayan, R.; Rinaldi, D.; Larburu, N.; Plassart, L.; Balor, S.; Bouyssié, D.; Lebaron, S.; Marcoux, J.; Gleizes, P.-E.; Plisson-Chastang, C. Good Vibrations: Structural Remodeling of Maturing Yeast Pre-40S Ribosomal Particles Followed by Cryo-Electron Microscopy. Molecules 2020, 25, 1125. https://doi.org/10.3390/molecules25051125
Shayan R, Rinaldi D, Larburu N, Plassart L, Balor S, Bouyssié D, Lebaron S, Marcoux J, Gleizes P-E, Plisson-Chastang C. Good Vibrations: Structural Remodeling of Maturing Yeast Pre-40S Ribosomal Particles Followed by Cryo-Electron Microscopy. Molecules. 2020; 25(5):1125. https://doi.org/10.3390/molecules25051125
Chicago/Turabian StyleShayan, Ramtin, Dana Rinaldi, Natacha Larburu, Laura Plassart, Stéphanie Balor, David Bouyssié, Simon Lebaron, Julien Marcoux, Pierre-Emmanuel Gleizes, and Célia Plisson-Chastang. 2020. "Good Vibrations: Structural Remodeling of Maturing Yeast Pre-40S Ribosomal Particles Followed by Cryo-Electron Microscopy" Molecules 25, no. 5: 1125. https://doi.org/10.3390/molecules25051125