https://www.cell.com/molecular-cell/pdf/S1097-2765(13)00326-2.pdf

eIF5A promotes translation of polyproline motifs.

eIF5A Functions Globally in Translation Elongation and Termination

https://www.cell.com/molecular-cell/pdf/S1097-2765(16)00009-5.pdf

Synonymous Codons Direct Cotranslational Folding toward Different Protein Conformations

Genetic decoding is not 'frozen' as was earlier thought, but dynamic. One facet of this is frameshifting that often results in synthesis of a C-terminal region encoded by a new frame. Ribosomal frameshifting is utilized for the synthesis of additional products, for regulatory purposes and for translational 'correction' of problem or 'savior' indels. Utilization for synthesis of additional products occurs prominently in the decoding of mobile chromosomal element and viral genomes. One class of regulatory frameshifting of stable chromosomal genes governs cellular polyamine levels from yeasts to humans. In many cases of productively utilized frameshifting, the proportion of ribosomes that frameshift at a shift-prone site is enhanced by specific nascent peptide or mRNA context features. Such mRNA signals, which can be 5' or 3' of the shift site or both, can act by pairing with ribosomal RNA or as stem loops or pseudoknots even with one component being 4 kb 3' from the shift site. Transcriptional realignment at slippage-prone sequences also generates productively utilized products encoded trans-frame with respect to the genomic sequence. This too can be enhanced by nucleic acid structure. Together with dynamic codon redefinition, frameshifting is one of the forms of recoding that enriches gene expression.

/pdf/ribosomal-frameshifting-and-transcriptional-slippage-from-2dxbxuyood.pdf

Ribosomal frameshifting and transcriptional slippage: From genetic steganography and cryptography to adventitious use.

Owing to the degeneracy of the genetic code, a protein sequence can be encoded by many different synonymous mRNA coding sequences. Synonymous codon usage was once thought to be functionally neutral, but evidence now indicates it is shaped by evolutionary selection and affects other aspects of protein biogenesis beyond specifying the amino acid sequence of the protein. Synonymous rare codons, once thought to have only negative impacts on the speed and accuracy of translation, are now known to play an important role in diverse functions, including regulation of cotranslational folding, covalent modifications, secretion, and expression level. Mutations altering synonymous codon usage are linked to human diseases. However, much remains unknown about the molecular mechanisms connecting synonymous codon usage to efficient protein biogenesis and proper cell physiology. Here we review recent literature on the functional effects of codon usage, including bioinformatics approaches aimed at identifying general roles for synonymous codon usage.

Roles for Synonymous Codon Usage in Protein Biogenesis

Elongation factor P (EF-P) is a translation factor of unknown function that has been implicated in a great variety of cellular processes. Here, we show that EF-P prevents ribosome from stalling during synthesis of proteins containing consecutive prolines, such as PPG, PPP, or longer proline strings, in natural and engineered model proteins. EF-P promotes peptide-bond formation and stabilizes the peptidyl–transfer RNA in the catalytic center of the ribosome. EF-P is posttranslationally modified by a hydroxylated β-lysine attached to a lysine residue. The modification enhances the catalytic proficiency of the factor mainly by increasing its affinity to the ribosome. We propose that EF-P and its eukaryotic homolog, eIF5A, are essential for the synthesis of a subset of proteins containing proline stretches in all cells.

/pdf/ef-p-is-essential-for-rapid-synthesis-of-proteins-containing-1l1251pwj2.pdf

EF-P is essential for rapid synthesis of proteins containing consecutive proline residues

The peptide bond formation with the amino acid proline (Pro) on the ribosome is slow, resulting in translational stalling when several Pro have to be incorporated into the peptide. Stalling at poly-Pro motifs is alleviated by the elongation factor P (EF-P). Here we investigate why Pro is a poor substrate and how EF-P catalyzes the reaction. Linear free energy relationships of the reaction on the ribosome and in solution using 12 different Pro analogues suggest that the positioning of Pro-tRNA in the peptidyl transferase center is the major determinant for the slow reaction. With any Pro analogue tested, EF-P decreases the activation energy of the reaction by an almost uniform value of 2.5 kcal/mol. The main source of catalysis is the favorable entropy change brought about by EF-P. Thus, EF-P acts by entropic steering of Pro-tRNA toward a catalytically productive orientation in the peptidyl transferase center of the ribosome.

Entropic Contribution of Elongation Factor P to Proline Positioning at the Catalytic Center of the Ribosome.

The elongation phase of translation is promoted by three universal elongation factors, EF-Tu, EF-Ts, and EF-G in bacteria and their homologs in archaea and eukaryotes Recent findings demonstrate that the translation of a subset of mRNAs requires a fourth elongation factor, EF-P in bacteria or the homologs factors a/eIF5A in other kingdoms of life EF-P prevents the ribosome from stalling during the synthesis of proteins containing consecutive Pro residues, such as PPG, PPP, or longer Pro clusters The efficient and coordinated synthesis of such proteins is required for bacterial growth, motility, virulence, and stress response EF-P carries a unique post-translational modification, which contributes to its catalytic proficiency The modification enzymes, which are lacking in higher eukaryotes, provide attractive new targets for the development of new, highly specific antimicrobials

/pdf/elongation-factor-p-function-and-effects-on-bacterial-3o5s16fkie.pdf

Elongation factor P: Function and effects on bacterial fitness.

Human mitochondrial gene expression relies on the specific recognition and aminoacylation of mitochondrial tRNAs (mtRNAs) by nuclear-encoded mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs). Despite their essential role in cellular energy homeostasis, strong mutation pressure and genetic drift have led to an unparalleled sequence erosion of animal mtRNAs. The structural and functional consequences of this erosion are not understood. Here, we present cryo-EM structures of the human mitochondrial seryl-tRNA synthetase (mSerRS) in complex with mtRNASer(GCU). These structures reveal a unique mechanism of substrate recognition and aminoacylation. The mtRNASer(GCU) is highly degenerated, having lost the entire D-arm, tertiary core, and stable L-shaped fold that define canonical tRNAs. Instead, mtRNASer(GCU) evolved unique structural innovations, including a radically altered T-arm topology that serves as critical identity determinant in an unusual shape-selective readout mechanism by mSerRS. Our results provide a molecular framework to understand the principles of mito-nuclear co-evolution and specialized mechanisms of tRNA recognition in mammalian mitochondrial gene expression. 

/pdf/structural-basis-for-shape-selective-recognition-and-2186fcxc.pdf

Structural basis for shape-selective recognition and aminoacylation of a D-armless human mitochondrial tRNA

Understanding the assembly principles of biological macromolecular complexes remains a significant challenge, due to the complexity of the systems and the difficulties in developing experimental approaches. As a ribonucleoprotein complex, the ribosome serves as an ideal model system for the profiling of macromolecular complex assembly. In this work, we report an ensemble of large ribosomal subunit intermediate structures that accumulate during synthesis in a near-physiological and co-transcriptional in vitro reconstitution system. Thirteen pre-50S intermediate maps covering the whole assembly process were resolved using cryo-EM single particle analysis and heterogeneous subclassification. Segmentation of the set of density maps reveals that the 50S ribosome intermediates assemble based on fourteen cooperative assembly blocks, including the smallest assembly core reported up to now, which is composed of a 600-nucleotide-long folded rRNA and three ribosomal proteins. The cooperative blocks assemble onto the assembly core following a defined set of dependencies, revealing the parallel assembly pathways at both early and late assembly stages of the 50S subunit.

/pdf/near-physiological-in-vitro-assembly-of-50s-ribosomes-ijg9w66f.pdf

Lili K. Doerfel

Papers

EF-P is essential for rapid synthesis of proteins containing consecutive proline residues

Entropic Contribution of Elongation Factor P to Proline Positioning at the Catalytic Center of the Ribosome.

Elongation factor P: Function and effects on bacterial fitness.

Structural basis for shape-selective recognition and aminoacylation of a D-armless human mitochondrial tRNA

Near-physiological in vitro assembly of 50S ribosomes involves parallel pathways