Ribosome Structure and the Mechanism of Translation

Valuable information on translation initiation is available from biochemical data and recently solved structures. We present a detailed description of current knowledge about the structure, function, and interactions of the individual components involved in bacterial translation initiation. The first section describes the ribosomal features relevant to the initiation process. Subsequent sections describe the structure, function, and interactions of the mRNA, the initiator tRNA, and the initiation factors IF1, IF2, and IF3. Finally, we provide an overview of mechanisms of regulation of the translation initiation event. Translation occurs on ribonucleoprotein complexes called ribosomes. The ribosome is composed of a large subunit and a small subunit that hold the activities of peptidyltransfer and decode the triplet code of the mRNA, respectively. Translation initiation is promoted by IF1, IF2, and IF3, which mediate base pairing of the initiator tRNA anticodon to the mRNA initiation codon located in the ribosomal P-site. The mechanism of translation initiation differs for canonical and leaderless mRNAs, since the latter is dependent on the relative level of the initiation factors. Regulation of translation occurs primarily in the initiation phase. Secondary structures at the mRNA ribosomal binding site (RBS) inhibit translation initiation. The accessibility of the RBS is regulated by temperature and binding of small metabolites, proteins, or antisense RNAs. The future challenge is to obtain atomic-resolution structures of complete initiation complexes in order to understand the mechanism of translation initiation in molecular detail.

Initiation of Protein Synthesis in Bacteria

The small ribosomal subunit is responsible for the decoding of genetic information and plays a key role in the initiation of protein synthesis. We analyzed by X-ray crystallography the structures of three different complexes of the small ribosomal subunit of Thermus thermophilus with the A-site inhibitor tetracycline, the universal initiation inhibitor edeine and the C-terminal domain of the translation initiation factor IF3. The crystal structure analysis of the complex with tetracycline revealed the functionally important site responsible for the blockage of the A-site. Five additional tetracycline sites resolve most of the controversial biochemical data on the location of tetracycline. The interaction of edeine with the small subunit indicates its role in inhibiting initiation and shows its involvement with P-site tRNA. The location of the C-terminal domain of IF3, at the solvent side of the platform, sheds light on the formation of the initiation complex, and implies that the anti-association activity of IF3 is due to its influence on the conformational dynamics of the small ribosomal subunit.

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Crystal structures of complexes of the small ribosomal subunit with tetracycline, edeine and IF3

Eukaryotic translation initiation factors and regulators

Translation, the process of mRNA-encoded protein synthesis, requires a complex apparatus, composed of the ribosome, tRNAs and additional protein factors, including aminoacyl tRNA synthetases. The ribosome provides the platform for proper assembly of mRNA, tRNAs and protein factors and carries the peptidyl-transferase activity. It consists of small and large subunits. The ribosomes are ribonucleoprotein particles with a ribosomal RNA core, to which multiple ribosomal proteins are bound. The sequence and structure of ribosomal RNAs, tRNAs, some of the ribosomal proteins and some of the additional protein factors are conserved in all kingdoms, underlying the common origin of the translation apparatus. Translation can be subdivided into several steps: initiation, elongation, termination and recycling. Of these, initiation is the most complex and the most divergent among the different kingdoms of life. A great amount of new structural, biochemical and genetic information on translation initiation has been accumulated in recent years, which led to the realization that initiation also shows a great degree of conservation throughout evolution. In this review, we summarize the available structural and functional data on translation initiation in the context of evolution, drawing parallels between eubacteria, archaea, and eukaryotes. We will start with an overview of the ribosome structure and of translation in general, placing emphasis on factors and processes with relevance to initiation. The major steps in initiation and the factors involved will be described, followed by discussion of the structure and function of the individual initiation factors throughout evolution. We will conclude with a summary of the available information on the kinetic and thermodynamic aspects of translation initiation.

https://www.cambridge.org/core/services/aop-cambridge-core/content/view/S0033583505004026

Translation initiation: structures, mechanisms and evolution.

Solution structure of the ribosome-binding domain of E. coli translation initiation factor IF3. Homology with the U1A protein of the eukaryotic spliceosome.

Heteronuclear NMR studies of E. coli translation initiation factor IF3. Evidence that the inter-domain region is disordered in solution.

Initiation factor IF3 from Escherichia coli is composed of two domains connected by a hydrophilic peptide. In this study, the N-terminal domain (residues 7–83) has been overexpiessed, 15N labelled and purified. NMR assignments for this domain have been obtained by two-dimensional and three-dimensional heteronuclear and homonuclear spectroscopy. Using distance geometry and simulated annealing, a three-dimensional solution structure was calculated using 506 NOE and 56 dihedral angle restraints. The resulting structure is composed of a five-stranded antiparallel β sheet surrounded by two α helices. Since the heteronuclear 1H-15N correlation spectrum of the N-terminal domain of IF3 is an almost exact subset of that of the native protein, the assignments obtained and the structure calculated should be directly transposable to the full-length protein.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1995.0395n.x

Carlos Garcia

Papers

Solution structure of the ribosome-binding domain of E. coli translation initiation factor IF3. Homology with the U1A protein of the eukaryotic spliceosome.

Heteronuclear NMR studies of E. coli translation initiation factor IF3. Evidence that the inter-domain region is disordered in solution.

1H and 15N resonance assignments and structure of the N-terminal domain of Escherichia coli initiation factor 3.

The N-terminal half of initiation factor IF3 is folded as a stable independent domain.