Entropic Contribution of Elongation Factor P to Proline Positioning at the Catalytic Center of the Ribosome.
14 Oct 2015-Journal of the American Chemical Society (American Chemical Society)-Vol. 137, Iss: 40, pp 12997-13006
TL;DR: Why Pro is a poor substrate and how EF-P catalyzes the reaction are investigated, which suggests that the positioning of Pro-tRNA in the peptidyl transferase center is the major determinant for the slow reaction.
Abstract: 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.
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TL;DR: Together with dynamic codon redefinition, frameshifting is one of the forms of recoding that enriches gene expression and generates productively utilized products encoded trans-frame with respect to the genomic sequence.
Abstract: 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.
260 citations
Cites background from "Entropic Contribution of Elongation..."
...Hypusine, and different modifications on the bacterial counterpart of eEF5, EF-P, likely evolved for the stabilization of the CCA-end of P-site tRNA (470,471) [Importance of CCA characteristics are shown by tRNA with CCA mutated to GCA or ACA causing frameshifting at a specific sequence and this being enhanced by mutants of hrpA (440,472) that encodes an RNA helicase (473)....
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TL;DR: The aim of this article is to show the link between ribosome structure, dynamics, and function.
Abstract: This review summarizes our current understanding of translation in prokaryotes, focusing on the mechanistic and structural aspects of each phase of translation: initiation, elongation, termination, and ribosome recycling. The assembly of the initiation complex provides multiple checkpoints for messenger RNA (mRNA) and start-site selection. Correct codon-anticodon interaction during the decoding phase of elongation results in major conformational changes of the small ribosomal subunit and shapes the reaction pathway of guanosine triphosphate (GTP) hydrolysis. The ribosome orchestrates proton transfer during peptide bond formation, but requires the help of elongation factor P (EF-P) when two or more consecutive Pro residues are to be incorporated. Understanding the choreography of transfer RNA (tRNA) and mRNA movements during translocation helps to place the available structures of translocation intermediates onto the time axis of the reaction pathway. The nascent protein begins to fold cotranslationally, in the constrained space of the polypeptide exit tunnel of the ribosome. When a stop codon is reached at the end of the coding sequence, the ribosome, assisted by termination factors, hydrolyzes the ester bond of the peptidyl-tRNA, thereby releasing the nascent protein. Following termination, the ribosome is dissociated into subunits and recycled into another round of initiation. At each step of translation, the ribosome undergoes dynamic fluctuations between different conformation states. The aim of this article is to show the link between ribosome structure, dynamics, and function.
164 citations
Additional excerpts
...4) (Doerfel et al. 2015; Huter et al. 2017)....
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TL;DR: The experimental evidence on how the ribosome can alter translational efficiencies of mRNA at the initiation and elongation stages and how translation velocity affects protein folding are summarized to argue in favour of a new understanding of translation control as a hub that links mRNA homeostasis to production and quality control of proteins in the cell.
Abstract: The cellular proteome is shaped by the combined activities of the gene expression and quality control machineries. While transcription plays an undoubtedly important role, in recent years also translation emerged as a key step that defines the composition and quality of the proteome and the functional activity of proteins in the cell. Among the different post-transcriptional control mechanisms, translation initiation and elongation provide multiple checkpoints that can affect translational efficiency. A multitude of specific signals in mRNAs can determine the frequency of translation initiation, choice of the open reading frame, global and local elongation velocities, and the folding of the emerging protein. In addition to specific signatures in the mRNAs, also variations in the global pools of translation components, including ribosomes, tRNAs, mRNAs, and translation factors can alter translational efficiencies. The cellular outcomes of phenomena such as mRNA codon bias are sometimes difficult to understand due to the staggering complexity of covariates that affect codon usage, translation, and protein folding. Here we summarize the experimental evidence on how the ribosome-together with the other components of the translational machinery-can alter translational efficiencies of mRNA at the initiation and elongation stages and how translation velocity affects protein folding. We seek to explain these findings in the context of mechanistic work on the ribosome. The results argue in favour of a new understanding of translation control as a hub that links mRNA homeostasis to production and quality control of proteins in the cell.
148 citations
Cites background from "Entropic Contribution of Elongation..."
...In comparison, the electronic properties or the ability to undergo cis-trans isomerization are less important.(109) In some cases, Pro-induced ribosome stalling leads to tmRNA-mediated peptide tagging and degradation by the trans-translation mechanism....
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TL;DR: It is seen that the balance between the basic steps in elongation and the less common recoding events is determined by the kinetics of the different processes as well as by specific sequence determinants.
Abstract: In this review, we highlight the current understanding of translation elongation and recoding in eukaryotes. In addition to providing an overview of the process, recent advances in our understanding of the role of the factor eIF5A in both translation elongation and termination are discussed. We also highlight mechanisms of translation recoding with a focus on ribosomal frameshifting during elongation. We see that the balance between the basic steps in elongation and the less common recoding events is determined by the kinetics of the different processes as well as by specific sequence determinants.
134 citations
TL;DR: When ribosomes are arrested during elongation or termination, th... as discussed by the authors showed that the ribosome can be killed during the process of polypeptide initiation, elongation, termination, and recycling.
Abstract: Ribosomes translate genetic information into polypeptides in several basic steps: initiation, elongation, termination and recycling. When ribosomes are arrested during elongation or termination, th...
129 citations
References
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TL;DR: These mutants—the ‘Keio collection’—provide a new resource not only for systematic analyses of unknown gene functions and gene regulatory networks but also for genome‐wide testing of mutational effects in a common strain background, E. coli K‐12 BW25113.
Abstract: We have systematically made a set of precisely defined, single-gene deletions of all nonessential genes in Escherichia coli K-12. Open-reading frame coding regions were replaced with a kanamycin cassette flanked by FLP recognition target sites by using a one-step method for inactivation of chromosomal genes and primers designed to create in-frame deletions upon excision of the resistance cassette. Of 4288 genes targeted, mutants were obtained for 3985. To alleviate problems encountered in high-throughput studies, two independent mutants were saved for every deleted gene. These mutants-the 'Keio collection'-provide a new resource not only for systematic analyses of unknown gene functions and gene regulatory networks but also for genome-wide testing of mutational effects in a common strain background, E. coli K-12 BW25113. We were unable to disrupt 303 genes, including 37 of unknown function, which are candidates for essential genes. Distribution is being handled via GenoBase (http://ecoli.aist-nara.ac.jp/).
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Abstract: Collagen is the most abundant protein in animals. This fibrous, structural protein comprises a right-handed bundle of three parallel, left-handed polyproline II-type helices. Much progress has been made in elucidating the structure of collagen triple helices and the physicochemical basis for their stability. New evidence demonstrates that stereoelectronic effects and preorganization play a key role in that stability. The fibrillar structure of type I collagen—the prototypical collagen fibril—has been revealed in detail. Artificial collagen fibrils that display some properties of natural collagen fibrils are now accessible using chemical synthesis and self-assembly. A rapidly emerging understanding of the mechanical and structural properties of native collagen fibrils will guide further development of artificial collagenous materials for biomedicine and nanotechnology.
2,742 citations
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TL;DR: The three-dimensional structure of proteins chemical catalysis the basic equations of enzyme kinetics measurement and magnitude of enzymatic rate constants the pH dependence of enzyme catalysis practical kinetics detection of intermediaries in reactions by kinetics stereochemistry of enzymes reactions active-site-directed and enzyme-activated irreversible inhibitors - affinity labels and suicide inhibitors conformational change, allosteric regulation, motors and work forces between molecules, and enzymesubstrate binding energies enzyme-substrate complementarity and the use of binding energy in catalysis specificity and editing mechanisms recombinant DNA technology case studies of enzyme
Abstract: The three-dimensional structure of proteins chemical catalysis the basic equations of enzyme kinetics measurement and magnitude of enzymatic rate constants the pH dependence of enzyme catalysis practical kinetics detection of intermediaries in reactions by kinetics stereochemistry of enzymic reactions active-site-directed and enzyme-activated irreversible inhibitors - affinity labels and suicide inhibitors conformational change, allosteric regulation, motors and work forces between molecules, and enzyme-substrate binding energies enzyme-substrate complementarity and the use of binding energy in catalysis specificity and editing mechanisms recombinant DNA technology case studies of enzyme structure and mechanism protein engineering protein stability kinetics of protein folding folding pathways and energy landscapes.
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