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Review
. 2017 Mar 19;372(1716):20160181.
doi: 10.1098/rstb.2016.0181.

Structure and Dynamics of Bacterial Ribosome Biogenesis

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Free PMC article
Review

Structure and Dynamics of Bacterial Ribosome Biogenesis

Joseph H Davis et al. Philos Trans R Soc Lond B Biol Sci. .
Free PMC article

Abstract

Bacterial ribosome biogenesis has been an active area of research for more than 30 years and has served as a test-bed for the development of new biochemical, biophysical and structural techniques to understand macromolecular assembly generally. Recent work inspecting the process in vivo has advanced our understanding of the role of ribosome biogenesis factors, the co-transcriptional nature of assembly, the kinetics of the process under sub-optimal conditions, and the rRNA folding and ribosome protein binding pathways. Additionally, new structural work enabled by single-particle electron microscopy has helped to connect in vitro ribosomal protein binding maps to the underlying RNA. This review summarizes the state of these in vivo studies, provides a kinetic model for ribosome assembly under sub-optimal conditions, and describes a framework to compare newly emerging assembly intermediate structures.This article is part of the themed issue 'Perspectives on the ribosome'.

Keywords: RNA folding; assembly factors; cryo-electron microscopy; macromolecular assembly; quantitative mass spectrometry; ribosome assembly.

Figures

Figure 1.
Figure 1.
Co-transcriptional assembly of ribosomes in the E. coli ribosomal RNA operon. The negative-stain electron micrograph of fixed chromatin shows the high transcriptional level of the ribosomal operon, where over a hundred RNAP molecules can be seen over the 5.5 kb segment [13]. The organization of the rRNA genes is shown below, where the primary rRNA precursor transcript is processed by a series of endonucleolytic cleavage reactions to produce the mature 16S, 23S and 5S rRNAs. The series of increasingly long nascent transcripts can be seen progressing from left to right, with evidence for co-transcriptional binding of ribosomal proteins. In addition, co-transcriptional rRNA processing is observed, indicated by the arrow, which liberates a pre-30S ribonucleoprotein complex prior to initiating the transcription of the 23S rRNA gene.
Figure 2.
Figure 2.
Modelling the effects of perturbations on the populations of ribosome assembly intermediates. (a) A simple assembly map involving two primary binding proteins, P1 and P2, and two secondary binding proteins, P3 and P4, in analogy to the Nomura and Nierhaus maps. Binding of P3 depends on prior binding of P1, and binding of P4 depends on prior binding of P2. This leads to a class of early binding proteins in blue, and a class of late-binding proteins in yellow. (b) Perturbation of assembly by limitation of P2 binding results in apparent changes in the observed binding order of assembly. Accelerated proteins shown in green and delayed proteins shown in red. (c) An explicit model enumerating all possible intermediates, based on the assembly maps in (a,b), can be used to simulate the flux through the assembly pathways and the populations of all of the intermediates using the Gillespie algorithm for stochastic dynamics [49]. The RNA is schematically shown as a grid of binding sites, and the proteins as circles. Icons are coloured by abundance according to the colour bar. In the permissive scheme, all protein-binding rate constants are the same. In the restrictive scheme, the rate constant for all steps involving P2 binding are reduced fivefold (dashed arrows). (d) The total population of each protein summed across all intermediates is shown for the permissive condition (black) and the restrictive condition (grey). These values correspond to the protein levels that would be measured by quantitative mass spectrometry. (e) Simulated pulse-labelling kinetics of the mature particle under permissive (blue) or restrictive (red) conditions. A no-intermediate reference is also shown (black).
Figure 3.
Figure 3.
Cooperative folding blocks of the bacterial large (LSU) and small (SSU) ribosomal subunits. (a) Heatmap of median folding block occupancy in various published SSU assembly intermediate structures. Blocks 1–4 were derived from hierarchical clustering of the calculated occupancy for each r-protein or rRNA helix across the set of available SSU assembly intermediate structures (electronic supplementary material, figure S1). Structures are labelled according to table 1. (b) 16S secondary structure coloured and labelled by domain. Folding blocks as defined by hierarchical clustering in the electronic supplementary material, figure S1, are outlined and coloured according to (a). (c) SSU structure model (PDB: 4ybb) with blocks labelled and coloured according to (a). (d) Heatmap of median folding block occupancy in various published LSU assembly intermediate structures. Blocks 1–6 were derived from hierarchical clustering as in (a) using the set of available LSU assembly intermediate structures (electronic supplementary material, figure S2) and are labelled according to table 1. (e) 23S secondary structure coloured and labelled by domain and folding blocks as in (b). (f) LSU structure model (PDB: 4ybb) with blocks labelled and coloured according to (d).

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