Final touches and quality control on the assembly of the eukaryotic ribosome

Aida Razi; Joaquin Ortega

doi:10.15252/embj.201796722

Eukaryotic ribosome assembly, transport and quality control

Nature Structural & Molecular Biology ◽

10.1038/nsmb.3454 ◽

2017 ◽

Vol 24 (9) ◽

pp. 689-699 ◽

Cited By ~ 97

Author(s):

Cohue Peña ◽

Ed Hurt ◽

Vikram Govind Panse

Keyword(s):

Quality Control ◽

Ribosome Assembly ◽

Eukaryotic Ribosome

Download Full-text

The GTPase Nog1 co-ordinates the assembly, maturation and quality control of distant ribosomal functional centers

eLife ◽

10.7554/elife.52474 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 7

Author(s):

Purnima Klingauf-Nerurkar ◽

Ludovic C Gillet ◽

Daniela Portugal-Calisto ◽

Michaela Oborská-Oplová ◽

Martin Jäger ◽

...

Keyword(s):

Quality Control ◽

Protein Folding ◽

Atpase Activity ◽

Ribosomal Protein ◽

Peptidyl Transferase ◽

Peptidyl Transferase Center ◽

Eukaryotic Ribosome ◽

Ribosomal Stalk ◽

Exit Tunnel ◽

Ribosome Formation

Eukaryotic ribosome precursors acquire translation competence in the cytoplasm through stepwise release of bound assembly factors, and proofreading of their functional centers. In case of the pre-60S, these steps include removal of placeholders Rlp24, Arx1 and Mrt4 that prevent premature loading of the ribosomal protein eL24, the protein-folding machinery at the polypeptide exit tunnel (PET), and the ribosomal stalk, respectively. Here, we reveal that sequential ATPase and GTPase activities license release factors Rei1 and Yvh1 to trigger Arx1 and Mrt4 removal. Drg1-ATPase activity removes Rlp24 from the GTPase Nog1 on the pre-60S; consequently, the C-terminal tail of Nog1 is extracted from the PET. These events enable Rei1 to probe PET integrity and catalyze Arx1 release. Concomitantly, Nog1 eviction from the pre-60S permits peptidyl transferase center maturation, and allows Yvh1 to mediate Mrt4 release for stalk assembly. Thus, Nog1 co-ordinates the assembly, maturation and quality control of distant functional centers during ribosome formation.

Download Full-text

The GTPase Nog1 couples polypeptide exit tunnel quality control with ribosomal stalk assembly

10.1101/462333 ◽

2018 ◽

Cited By ~ 4

Author(s):

Purnima Nerurkar ◽

Ludovic Gillet ◽

Cohue Pena ◽

Olga T Schubert ◽

Martin Altvater ◽

...

Keyword(s):

Quality Control ◽

Protein Folding ◽

Atpase Activity ◽

Large Subunit ◽

Eukaryotic Ribosome ◽

Release Factors ◽

Assembly Factors ◽

Ribosomal Stalk ◽

Exit Tunnel ◽

Ribosome Formation

Eukaryotic ribosome precursors acquire translation competence in the cytoplasm through stepwise release of bound assembly factors, and proofreading of their functional centers. In case of the large subunit precursor (pre-60S), these essential steps include eviction of placeholders Arx1 and Mrt4 that prevent premature loading of the protein-folding machinery at the polypeptide exit tunnel (PET), and the ribosomal stalk, respectively. Here, we reveal that sequential ATPase and GTPase activities license release factors Rei1 and Yvh1 recruitment to the pre-60S in order to trigger Arx1 and Mrt4 removal. Drg1-ATPase activity extracts the C-terminal tail of Nog1 from the PET, enabling Rei1 to probe PET integrity, and then catalyze Arx1 release. Subsequently, GTPase hydrolysis stimulates Nog1 removal from the pre-60S, permitting Yvh1 to mediate Mrt4 release, and initiate ribosomal stalk assembly. Thus, Nog1 couples quality control and assembly of spatially distant functional centers during ribosome formation.

Download Full-text

The native structure of the eukaryotic ribosome

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100075890 ◽

1983 ◽

Vol 41 ◽

pp. 432-433

Author(s):

R.A. Milligan ◽

P.N.T. Unwin

Keyword(s):

Ribosomal Proteins ◽

Crystal Form ◽

Native Structure ◽

X Ray Diffraction ◽

Eukaryotic Ribosome ◽

Vitro Analysis ◽

In Vitro Analysis ◽

Resolution Structure ◽

Stable Unit

A detailed understanding of the mechanism of protein synthesis will ultimately depend on knowledge of the native structure of the ribosome. Towards this end we have investigated the low resolution structure of the eukaryotic ribosome embedded in frozen buffer, making use of a system in which the ribosomes crystallize naturally.The ribosomes in the cells of early chicken embryos form crystalline arrays when the embryos are cooled at 4°C. We have developed methods to isolate the stable unit of these arrays, the ribosome tetramer, and have determined conditions for the growth of two-dimensional crystals in vitro, Analysis of the proteins in the crystals by 2-D gel electrophoresis demonstrates the presence of all ribosomal proteins normally found in polysomes. There are in addition, four proteins which may facilitate crystallization. The crystals are built from two oppositely facing P4 layers and the predominant crystal form, accounting for >80% of the crystals, has the tetragonal space group P4212, X-ray diffraction of crystal pellets demonstrates that crystalline order extends to ~ 60Å.

Download Full-text

Structural Studies on the Eukaryotic Ribosome

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100180033 ◽

1990 ◽

Vol 48 (1) ◽

pp. 256-257

Author(s):

Adriana Verschoor ◽

Ronald Milligan ◽

Suman Srivastava ◽

Joachim Frank

Keyword(s):

Chick Embryo ◽

3D Reconstruction ◽

Single Particle ◽

Mass Density ◽

Particle Surface ◽

Uranyl Acetate ◽

Electron Microscopic ◽

Eukaryotic Ribosome ◽

Negative Stain ◽

Reconstruction Methods

We have studied the eukaryotic ribosome from two vertebrate species (rabbit reticulocyte and chick embryo ribosomes) in several different electron microscopic preparations (Fig. 1a-d), and we have applied image processing methods to two of the types of images. Reticulocyte ribosomes were examined in both negative stain (0.5% uranyl acetate, in a double-carbon preparation) and frozen hydrated preparation as single-particle specimens. In addition, chick embryo ribosomes in tetrameric and crystalline assemblies in frozen hydrated preparation have been examined. 2D averaging, multivariate statistical analysis, and classification methods have been applied to the negatively stained single-particle micrographs and the frozen hydrated tetramer micrographs to obtain statistically well defined projection images of the ribosome (Fig. 2a,c). 3D reconstruction methods, the random conical reconstruction scheme and weighted back projection, were applied to the negative-stain data, and several closely related reconstructions were obtained. The principal 3D reconstruction (Fig. 2b), which has a resolution of 3.7 nm according to the differential phase residual criterion, can be compared to the images of individual ribosomes in a 2D tetramer average (Fig. 2c) at a similar resolution, and a good agreement of the general morphology and of many of the characteristic features is seen.Both data sets show the ribosome in roughly the same ’view’ or orientation, with respect to the adsorptive surface in the electron microscopic preparation, as judged by the agreement in both the projected form and the distribution of characteristic density features. The negative-stain reconstruction reveals details of the ribosome morphology; the 2D frozen-hydrated average provides projection information on the native mass-density distribution within the structure. The 40S subunit appears to have an elongate core of higher density, while the 60S subunit shows a more complex pattern of dense features, comprising a rather globular core, locally extending close to the particle surface.

Download Full-text