Rate Of Chain Elongation Research Articles

Temperature is an underestimated parameter to regulate the start-up of enhanced carbon chain elongation

Temperature is an important environmental factor for optimized bioreactions. In this study, seven temperatures between 25 and 55 °C were undertaken to explore the product spectrums of carbon chain elongation in a mixed culture. Temperature was manifested to significantly affect the production rate, productivity, and selectivity of carbon chain elongation. 40 °C resulted in the highest caproate concentration (138.94 mmol-C/L) and the highest caproate selectivity up to 84.5 %. Combining microbial quantitative and diversity analysis, it was identified that the optimized temperature was owing to the competitive abilities of metabolically different species. The dominant bio-elongation Clostridium kluyveri preferred 35–45 °C. The competitive fermentative bacteria Clostridium magnum and Clostridium butyricum for acetate and butyrate anabolism preferred 25–35 °C, therefore, excluding lower temperature bio-elongation because of more acetate and butyrate conversion but not caproate. Metabolically diverse Pseudomonas sp was abundant at higher temperatures (50–55 °C) and longer duration, therefore, excluding thermophilic bio-elongation because of reduced caproate productivity.

Open Culture Ethanol-Based Chain Elongation to Form Medium Chain Branched Carboxylates and Alcohols.

Chain elongation fermentation allows for the synthesis of biobased chemicals from complex organic residue streams. To expand the product spectrum of chain elongation technology and its application range we investigated 1) how to increase selectivity towards branched chain elongation and 2) whether alternative branched carboxylates such as branched valerates can be used as electron acceptors. Elongation of isobutyrate elongation towards 4-methyl-pentanoate was achieved with a selectivity of 27% (of total products, based on carbon atoms) in a continuous system that operated under CO2 and acetate limited conditions. Increasing the CO2 load led to more in situ acetate formation that increased overall chain elongation rate but decreased the selectivity of branched chain elongation. A part of this acetate formation was related to direct ethanol oxidation that seemed to be thermodynamically coupled to hydrogenotrophic carboxylate reduction to corresponding alcohols. Several alcohols including isobutanol and n-hexanol were formed. The microbiome from the continuous reactor was also able to form small amounts of 5-methyl-hexanoate likely from 3-methyl-butanoate and ethanol as substrate in batch experiments. The highest achieved concentration of isoheptanoate was 6.4 ± 0.9 mM Carbon, or 118 ± 17 mg/L, which contributed for 7% to the total amount of products (based on carbon atoms). The formation of isoheptanoate was dependent on the isoform of branched valerate. With 3-methyl-butanoate as substrate 5-methylhexanoate was formed, whereas a racemic mixture of L/D 2-methyl-butanoate did not lead to an elongated product. When isobutyrate and isovalerate were added simultaneously as substrates there was a large preference for elongation of isobutyrate over isovalerate. Overall, this work showed that chain elongation microbiomes can be further adapted with supplement of branched-electron acceptors towards the formation of iso-caproate and iso-heptanoate as well as that longer chain alcohol formation can be stimulated.

Translation of Human Globin mRNA: Globin Synthesis in Cells Containing Hb Leiden

Most structurally abnormal hemoglobins are present in smaller amounts than HbA in the erythrocytes of heterozygous subjects. In the presence of a hemoglobinopathy, alpha and beta globin synthesis remains balanced with equal production of the two types of chains. In reticulocytes of subjects with Hb Leiden (beta 6 or 7 glu leads to 0) there is greater production of alpha than beta globin in vitro (beta/alpha = 0.67), and slightly more beta A is synthesized than beta Leiden (beta A/beta Leiden = 1.28). Differences in specific mRNA content, rates of initiation of chain synthesis, or rates of chain elongation could be responsible for such differential polypeptide synthesis. In the present study, the ribosomal assembly of beta A, beta Leiden, and alpha globin chains was examined in peripheral blood. The translation times of the three chains did not differ significantly (average times: beta A = 65.4 sec, beta Leiden = 70.8 sec, alpha = 53.5 sec). These results indicated that an altered rate of translation was not the source of the anomalous globin synthesis observed in vitro in cells containing Hb Leiden. The experiments suggested that the observed imbalance in alpha/beta production was due to either differential rates of initiation of globin chain synthesis or quantitative differences in the amounts of the specific mRNAs present in the cells.

Control of ribosome synthesis in bacteria: the important role of rRNA chain elongation rate.

Bacteria growth depends crucially on protein synthesis, which is limited by ribosome synthesis. Ribosomal RNA (rRNA) transcription is the rate-limiting step of ribosome synthesis. It is generally proposed that the transcriptional initiation rate of rRNA operon is the primary factor that controls the rRNA synthesis. In this study, we established a convenient GFP-based reporter approach for measuring the bacterial rRNA chain elongation rate. We showed that the rRNA chain elongation rate of Escherichia coli remains constant under nutrient limitation and chloramphenicol inhibition. In contrast, rRNA chain elongation rate decreases dramatically under low temperatures. Strikingly, we found that Vibrio natriegens, the fastest growing bacteria known, has a 50% higher rRNA chain elongation rate than E. coli, which contributes to its rapid ribosome synthesis. Our study demonstrates that rRNA chain elongation rate is another important factor that affects the bacterial ribosome synthesis capacity.

Length Specificity and Polymerization Mechanism of (1,3)-β-d-Glucan Synthase in Fungal Cell Wall Biosynthesis.

(1,3)-β-d-Glucan synthase (GS) catalyzes formation of the linear (1,3)-β-d-glucan in the fungal cell wall and is a target of clinically approved antifungal antibiotics. The catalytic subunit of GS, FKS protein, does not exhibit significant sequence homology to other glycosyltransferases, and thus, significant ambiguity about its catalytic mechanism remains. One of the major technical barriers in studying GS is the absence of activity assay methods that allow characterization of the lengths and amounts of (1,3)-β-d-glucan due to its poor solubility in water and organic solvents. Here, we report a successful development of a novel GS activity assay based on size-exclusion chromatography coupled with pulsed amperometric detection and radiation counting (SEC-PAD-RC), which allows for the simultaneous characterization of the amount and length of the polymer product. The assay revealed that the purified yeast GS produces glucan with a length of 6550 ± 760 mer, consistent with the reported degree of polymerization of (1,3)-β-d-glucan isolated from intact cells. Pre-steady state kinetic analysis revealed a highly efficient but rate-determining chain elongation rate of 51.5 ± 9.8 s-1, which represents the first observation of chain elongation by a nucleotide-sugar-dependent polysaccharide synthase. Coupling the SEC-PAD-RC method with substrate analogue mechanistic probes provided the first unambiguous evidence that GS catalyzes non-reducing end polymerization. On the basis of these observations, we propose a detailed model for the catalytic mechanism of GS. The approaches described here can be used to determine the mechanism of catalysis of other polysaccharide synthases.

Role of Polyubiquitin Chain Flexibility in the Catalytic Mechanism of Cullin-RING Ubiquitin Ligases.

Cullin-RING ubiquitin ligases are a diverse family of ubiquitin ligases that catalyze the synthesis of K48-linked polyubiquitin (polyUb) chains on a variety of substrates, ultimately leading to their degradation by the proteasome. The cullin-RING enzyme scaffold processively attaches a Ub molecule to the distal end of a growing chain up to lengths of eight Ub monomers. However, the molecular mechanism governing how chains of increasing size are built using a scaffold of largely fixed dimensions is not clear. We developed coarse-grained molecular dynamics simulations to describe the dependence of kcat for cullin-RING ligases on the length and flexibility of the K48-linked polyUb chain attached to the substrate protein, key factors that determine the rate of subsequent Ub attachment to the chain, and therefore, the ensuing biological outcomes of ubiquitination. The results suggest that a number of regulatory mechanisms may lead to variations in the rate of chain elongation for different cullin-RING ligases. Specifically, modulation of the distance between the target lysine and the phosphodegron sequence of the substrate, the distance between the substrate lysine and the active site cysteine of the Ub conjugation enzyme (E2) bound to the cullin-RING scaffold, and flexibility of the bound E2 can lead to significant differences in the processing of K48-linked chains on substrates, potentially leading to differences in biological outcomes.

Structural Dynamics of the Activation of Elongation Factor 2 Kinase by Ca2 +-Calmodulin

Eukaryotic elongation factor 2 kinase (eEF-2K), the only known calmodulin (CaM)-activated α-kinase, phosphorylates eukaryotic elongation factor 2 (eEF-2) on a specific threonine (Thr-56) diminishing its affinity for the ribosome and reducing the rate of nascent chain elongation during translation. Despite its critical cellular role, the precise mechanisms underlying the CaM-mediated activation of eEF-2K remain poorly defined. Here, employing a minimal eEF-2K construct (TR) that exhibits activity comparable to the wild-type enzyme and is fully activated by CaM in vitro and in cells, and using a variety of complimentary biophysical techniques in combination with computational modeling, we provide a structural mechanism by which CaM activates eEF-2K. Native mass analysis reveals that CaM, with two bound Ca2+ ions, forms a stoichiometric 1:1 complex with TR. Chemical crosslinking mass spectrometry and small-angle X-ray scattering measurements localize CaM near the N-lobe of the TR kinase domain and the spatially proximal C-terminal helical repeat. Hydrogen/deuterium exchange mass spectrometry and methyl NMR indicate that the conformational changes induced on TR by the engagement of CaM are not localized but are transmitted to remote regions that include the catalytic site and the functionally important phosphate binding pocket. The structural insights obtained from the present analyses, together with our previously published kinetics data, suggest that TR, and by inference, wild-type eEF-2K, upon engaging CaM undergoes a conformational transition resulting in a state that is primed to efficiently auto-phosphorylate on the primary activating T348 en route to full activation.

Model-based genome-wide determination of RNA chain elongation rates in Escherichia coli

Dynamics in the process of transcription are often simplified, yet they play an important role in transcript folding, translation into functional protein and DNA supercoiling. While the modulation of the speed of transcription of individual genes and its role in regulation and proper protein folding has been analyzed in depth, the functional relevance of differences in transcription speeds as well as the factors influencing it have not yet been determined on a genome-wide scale. Here we determined transcription speeds for the majority of E. coli genes based on experimental data. We find large differences in transcription speed between individual genes and a strong influence of both cellular location as well as the relative importance of genes for cellular function on transcription speeds. Investigating factors influencing transcription speeds we observe both codon composition as well as factors associated to DNA topology as most important factors influencing transcription speeds. Moreover, we show that differences in transcription speeds are sufficient to explain the timing of regulatory responses during environmental shifts and highlight the importance of the consideration of transcription speeds in the design of experiments measuring transcriptomic responses to perturbations.

Roles of Conserved Active Site Residues in the Ketosynthase Domain of an Assembly Line Polyketide Synthase.

Ketosynthase (KS) domains of assembly line polyketide synthases (PKSs) catalyze intermodular translocation of the growing polyketide chain as well as chain elongation via decarboxylative Claisen condensation. The mechanistic roles of ten conserved residues in the KS domain of Module 1 of the 6-deoxyerythronolide B synthase were interrogated via site-directed mutagenesis and extensive biochemical analysis. Although the C211A mutant at the KS active site exhibited no turnover activity, it was still a competent methylmalonyl-ACP decarboxylase. The H346A mutant exhibited reduced rates of both chain translocation and chain elongation, with a greater effect on the latter half-reaction. H384 contributed to methylmalonyl-ACP decarboxylation, whereas K379 promoted C-C bond formation. S315 played a role in coupling decarboxylation to C-C bond formation. These findings support a mechanism for the translocation and elongation half-reactions that provides a well-defined starting point for further analysis of the key chain-building domain in assembly line PKSs.

Optimum alcohol concentration for chain elongation in mixed-culture fermentation of cellulosic substrate.

Medium-chain fatty acids (MCFA, e.g., caproic, heptanoic, caprylic acid) are more valuable than short-chain fatty acids (SCFA, e.g., acetic, propionic, butyric, valeric acid). SCFAs are major products in methane-inhibited mixed-culture anaerobic fermentation. By feeding ethanol to the fermentor, MCFA formation is enhanced through chain elongation. Microorganisms such as Clostridium kluyveri elongate short-chain acids by combining them with alcohol. Very low ethanol concentration reduces chain elongation rates, whereas very high ethanol concentrations inhibit microorganisms. To maximize MCFA production, different ethanol concentrations were investigated in the mixed-culture fermentation of office paper and chicken manure. At 10 g/L ethanol concentration, 10 g/L MCFA was formed. High ethanol concentrations (above 40 g/L) inhibit microorganisms resulting in no chain elongation. For chain elongation, propanol was found to be more inhibitory than ethanol. The data suggest that MCFA production will increase by continuously extracting MCFA and maintaining 5-10 g/L ethanol concentration by periodic addition. Biotechnol. Bioeng. 2016;113: 2597-2604. © 2016 Wiley Periodicals, Inc.

Formyl-methionine as a degradation signal at the N-termini of bacterial proteins.

In bacteria, all nascent proteins bear the pretranslationally formed N-terminal formyl-methionine (fMet) residue. The fMet residue is cotranslationally deformylated by a ribosome-associated deformylase. The formylation of N-terminal Met in bacterial proteins is not strictly essential for either translation or cell viability. Moreover, protein synthesis by the cytosolic ribosomes of eukaryotes does not involve the formylation of N-terminal Met. What, then, is the main biological function of this metabolically costly, transient, and not strictly essential modification of N terminal Met, and why has Met formylation not been eliminated during bacterial evolution? One possibility is that the similarity of the formyl and acetyl groups, their identical locations in N terminally formylated (Nt formylated) and Nt-acetylated proteins, and the recently discovered proteolytic function of Nt-acetylation in eukaryotes might also signify a proteolytic role of Nt formylation in bacteria. We addressed this hypothesis about fMet based degradation signals, termed fMet/N-degrons, using specific E. coli mutants, pulse-chase degradation assays, and protein reporters whose deformylation was altered, through site-directed mutagenesis, to be either rapid or relatively slow. Our findings strongly suggest that the formylated N-terminal fMet can act as a degradation signal, largely a cotranslational one. One likely function of fMet/N-degrons is the control of protein quality. In bacteria, the rate of polypeptide chain elongation is nearly an order of magnitude higher than in eukaryotes. We suggest that the faster emergence of nascent proteins from bacterial ribosomes is one mechanistic and evolutionary reason for the pretranslational design of bacterial fMet/N degrons, in contrast to the cotranslational design of analogous Ac/N degrons in eukaryotes.

Inhibitors of polyhydroxyalkanoate (PHA) synthases: synthesis, molecular docking, and implications.

Polyhydroxyalkanoate (PHA) synthases (PhaCs) catalyze the formation of biodegradable PHAs that are considered to be ideal alternatives to non-biodegradable synthetic plastics. However, study of PhaCs has been challenging because the rate of PHA chain elongation is much faster than that of initiation. This difficulty, along with lack of a crystal structure, has become the main hurdle to understanding and engineering PhaCs for economical PHA production. Here we report the synthesis of two carbadethia CoA analogues--sT-CH2-CoA (26 a) and sTet-CH2-CoA (26 b)--as well as sT-aldehyde (saturated trimer aldehyde, 29), as new PhaC inhibitors. Study of these analogues with PhaECAv revealed that 26 a/b and 29 are competitive and mixed inhibitors, respectively. Both the CoA moiety and extension of PHA chain will increase binding affinity; this is consistent with our docking study. Estimation of the Kic values of 26 a and 26 b predicts that a CoA analogue incorporating an octameric hydroxybutanoate (HB) chain might facilitate the formation of a kinetically well-behaved synthase.

Nanosecond Dynamics of Calmodulin and Ribosome‐Bound Nascent Chains Studied by Time‐Resolved Fluorescence Anisotropy

We report a time-resolved fluorescence anisotropy study of ribosome-bound nascent chains (RNCs) of calmodulin (CaM), a prototypical member of the EF-hand family of calcium-sensing proteins. As shown in numerous studies, in vitro protein refolding can differ substantially from biosynthetic protein folding, which takes place cotranslationally and depends on the rate of polypeptide chain elongation. A challenge in this respect is to characterize the adopted conformations of nascent chains before their release from the ribosome. CaM RNCs (full-length, half-length, and first EF-hand only) were synthesized in vitro. All constructs contained a tetracysteine motif site-specifically incorporated in the first N-terminal helix; this motif is known to react with FlAsH, a biarsenic fluorescein derivative. As the dye is rotationally locked to this helix, we characterized the structural properties and folding states of polypeptide chains tethered to ribosomes and compared these with released chains. Importantly, we observed decelerated tumbling motions of ribosome-tethered and partially folded nascent chains, compared to released chains. This indicates a pronounced interaction between nascent chains and the ribosome surface, and might reflect chaperone activity of the ribosome.

A SnapShot of Ubiquitin Chain Elongation: LYSINE 48-TETRA-UBIQUITIN SLOWS DOWN UBIQUITINATION

We have explored the mechanisms of polyubiquitin chain assembly with reconstituted ubiquitination of IκBα and β-catenin by the Skp1-cullin 1-βTrCP F-box protein (SCF(βTrCP)) E3 ubiquitin (Ub) ligase complex. Competition experiments revealed that SCF(βTrCP) formed a complex with IκBα and that the Nedd8 modified E3-substrate platform engaged in dynamic interactions with the Cdc34 E2 Ub conjugating enzyme for chain elongation. Using "elongation intermediates" containing β-catenin linked with Ub chains of defined length, it was observed that a Lys-48-Ub chain of a length greater than four, but not its Lys-63 linkage counterparts, slowed the rate of additional Ub conjugation. Thus, the Ub chain length and linkage impact kinetic rates of chain elongation. Given that Lys-48-tetra-Ub is packed into compact conformations due to extensive intrachain interactions between Ub subunits, this topology may limit the accessibility of SCF(βTrCP)/Cdc34 to the distal Ub Lys-48 and result in slowed elongation.

Thermodynamic Modeling of Variations in the Rate of RNA Chain Elongation of E. coli rrn Operons

Previous electron-microscopic imaging has shown high RNA polymerase occupation densities in the 16S and 23S encoding regions and low occupation densities in the noncoding leader, spacer, and trailer regions of the rRNA (rrn) operons in E. coli. This indicates slower transcript elongation within the coding regions and faster elongation within the noncoding regions of the operon. Inactivation of four of the seven rrn operons increases the transcript initiation frequency at the promoters of the three intact operons and reduces the time for RNA polymerase to traverse the operon. We have used the DNA sequence-dependent standard free energy variation of the transcription complex to model the experimentally observed changes in the elongation rate along the rrnB operon. We also model the stimulation of the average transcription rate over the whole operon by increasing rate of transcript initiation. Monte Carlo simulations, taking into account initiation of transcription, translocation, and backward and forward tracking of RNA polymerase, partially reproduce the observed transcript elongation rate variations along the rrn operon and fully account for the increased average rate in response to increased frequency of transcript initiation.

A single‐molecule approach to DNA replication in Escherichia coli cells demonstrated that DNA polymerase III is a major determinant of fork speed

The replisome catalyses DNA synthesis at a DNA replication fork. The molecular behaviour of the individual replisomes, and therefore the dynamics of replication fork movements, in growing Escherichia coli cells remains unknown. DNA combing enables a single-molecule approach to measuring the speed of replication fork progression in cells pulse-labelled with thymidine analogues. We constructed a new thymidine-requiring strain, eCOMB (E. coli for combing), that rapidly and sufficiently incorporates the analogues into newly synthesized DNA chains for the DNA-combing method. In combing experiments with eCOMB, we found the speed of most replication forks in the cells to be within the narrow range of 550-750 nt s(-1) and the average speed to be 653 ± 9 nt s(-1) (± SEM). We also found the average speed of the replication fork to be only 264 ± 9 nt s(-1) in a dnaE173-eCOMB strain producing a mutant-type of the replicative DNA polymerase III (Pol III) with a chain elongation rate (300 nt s(-1) ) much lower than that of the wild-type Pol III (900 nt s(-1) ). This indicates that the speed of chain elongation by Pol III is a major determinant of replication fork speed in E. coli cells.

Coupled activation and degradation of eEF2K regulates protein synthesis in response to genotoxic stress.

The kinase eEF2K [eukaryotic elongation factor 2 (eEF2) kinase] controls the rate of peptide chain elongation by phosphorylating eEF2, the protein that mediates the movement of the ribosome along the mRNA by promoting translocation of the transfer RNA from the A to the P site in the ribosome. eEF2K-mediated phosphorylation of eEF2 on threonine 56 (Thr⁵⁶) decreases its affinity for the ribosome, thereby inhibiting elongation. Here, we show that in response to genotoxic stress, eEF2K was activated by AMPK (adenosine monophosphate-activated protein kinase)-mediated phosphorylation on serine 398. Activated eEF2K phosphorylated eEF2 and induced a temporary ribosomal slowdown at the stage of elongation. Subsequently, during DNA damage checkpoint silencing, a process required to allow cell cycle reentry, eEF2K was degraded by the ubiquitin-proteasome system through the ubiquitin ligase SCF(βTrCP) (Skp1-Cul1-F-box protein, β-transducin repeat-containing protein) to enable rapid resumption of translation elongation. This event required autophosphorylation of eEF2K on a canonical βTrCP-binding domain. The inability to degrade eEF2K during checkpoint silencing caused sustained phosphorylation of eEF2 on Thr⁵⁶ and delayed the resumption of translation elongation. Our study therefore establishes a link between DNA damage signaling and translation elongation.

Role of a Conserved Arginine Residue in Linkers between the Ketosynthase and Acyltransferase Domains of Multimodular Polyketide Synthases

The role of interdomain linkers in modular polyketide synthases is poorly understood. Analysis of the 6-deoxyerythronolide B synthase (DEBS) has yielded a model in which chain elongation is governed by interactions between the acyl carrier protein domain and the ketosynthase domain plus an adjacent linker. Alanine scanning mutagenesis of the conserved residues of this linker in DEBS module 3 led to the identification of the R513A mutant with a markedly reduced rate of chain elongation. Limited proteolysis supported a structural role for this Arg. Our findings highlight the importance of domain-linker interactions in assembly line polyketide biosynthesis.

Structure and function of a HECT domain ubiquitin‐binding site

The Rsp5 ubiquitin ligase contains a non-covalent binding site for ubiquitin within the amino-terminal lobe (N-lobe) of the HECT domain, and the X-ray crystal structure of the HECT-ubiquitin complex has been determined. Hydrophobic patch residues of ubiquitin (L8, I44, V70) were crucial for interaction with Rsp5, and amino-acid alterations at the Rsp5-binding interface resulted in defects in polyubiquitination. Our results support a model in which the N-lobe-binding site acts to localize and orient the distal end of the ubiquitin chain to promote conjugation of the next ubiquitin molecule.

Chronic improvement of amino acid nutrition stimulates initiation of global messenger ribonucleic acid translation in tissues of sheep without affecting protein elongation1

Initiation of mRNA translation and elongation of the polypeptide chain are 2 regulated processes responsible for the short-term postprandial acceleration of protein synthesis in animal tissues. It is known that a chronic increase in the absorptive supply of AA stimulates protein synthesis in ruminant animals, but effects on translation initiation and elongation are unknown. To determine whether initiation or elongation phases of global mRNA translation are affected by chronic elevation of AA supply, 24 ewe lambs of 25.9 +/- 2.5 kg of BW were randomly allocated to 4 treatment groups of 6 lambs each. All lambs received a basal diet of barley and hay at 1.2 times maintenance ME intake. Treatments were an intravenous (i.v.) saline infusion as a control, i.v. infusion of 6 essential AA (EAA; Arg, Lys, His, Thr, Met, Cys) for 10 d, i.v. infusion of the same EAA excluding Met and Cys (EAA-SAA) for 10 d, and an oral drench of fishmeal twice daily for 17 d. Fishmeal supplementation supplied an extra 719 mg of N x kg(-0.75) x d(-1) and N retention was increased 519 mg x kg(-0.75) x d(-1) over the control. The EAA treatment supplied an extra 343 mg of N x kg(-0.75) x d(-1) directly into the blood, and N balance was increased by 268 mg x kg(-0.75) x d(-1). Deletion of Met plus Cys from EAA had no effect on N balance. The results indicate that Met plus Cys did not limit body protein gain on the basal diet alone or the basal diet plus 6 AA. Protein fractional synthesis rates in liver, duodenum, skin, rumen, semimembranosus, and LM were measured by a flooding dose procedure using L-[ring-2,6-(3)H]-Phe. Ribosome transit times were estimated from the ratio of nascent to total protein-bound radioactivities. Fishmeal and EAA treatments had no effect on RNA, DNA, or protein contents of tissues, but fractional synthesis rate, translational efficiency, and concentrations of active ribosomes were consistently elevated. Ribosome transit time was not affected by long-term AA supply. We conclude that the chronic stimulation of protein synthesis by long-term i.v. infusion of EAA or supplementation with an undegradable protein source is brought about by an improvement in the rate of initiation of mRNA translation with no change in the rate of polypeptide chain elongation.