Other Biochemistry, Biophysics, and Structural Biology Commons^™

Editing Of Misaminoacylated Trna Controls The Sensitivity Of Amino Acid Stress Responses In Saccharomyces Cerevisiae, Kyle Mohler, Rebecca Mann, Tammy J. Bullwinkle, Kyle W. Hopkins, Lin Hwang, Noah M. Reynolds, Brandon Gassaway, Hans-Rudolph Aerni, Jesse Rinehart, Michael Polymenis, Kym F. Faull, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Amino acid starvation activates the protein kinase Gcn2p, leading to changes in gene expression and translation. Gcn2p is activated by deacylated tRNA, which accumulates when tRNA aminoacylation is limited by lack of substrates or inhibition of synthesis. Pairing of amino acids and deacylated tRNAs is catalyzed by aminoacyl-tRNA synthetases, which use quality control pathways to maintain substrate specificity. Phenylalanyl-tRNA synthetase (PheRS) maintains specificity via an editing pathway that targets non-cognate Tyr-tRNA^Phe. While the primary role of aaRS editing is to prevent misaminoacylation, we demonstrate editing of misaminoacylated tRNA is also required for detection of amino acid starvation by …

An Assessment Of Potential False Positive E.Coli Pyroprints In The Cplop Database, Skyler A. Gordon

Master's Theses

The genetic information found in each species of organism is unique, and can be used as a tool to differentiate at the molecular level. This has caused rapid genotyping methods to become the cornerstone of a new area of research dependent on reading the genome as a form of identification. One of these specific identification methods, known as pyroprinting, relies on the small variation of DNA sequences within the same species to develop a unique, reproducible fingerprint. By simultaneously pyrosequencing multiple polymorphic loci within the ribosomal operons known as the intergenic transcribed spacers, a reproducible output is obtained, known as …

Quality Control By Isoleucyl-Trna Synthetase Of Bacillus Subtilis Is Required For Efficient Sporulation, Elizabeth Kermgard, Zhou Yang, Annika-Marisa Michel, Rachel Simari, Jacqueline Wong, Michael Ibba, Beth A. Lazazzera

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Isoleucyl-tRNA synthetase (IleRS) is an aminoacyl-tRNA synthetase whose essential function is to aminoacylate tRNA^Ile with isoleucine. Like some other aminoacyl-tRNA synthetases, IleRS can mischarge tRNA^Ile and correct this misacylation through a separate post-transfer editing function. To explore the biological significance of this editing function, we created a ileS(T233P) mutant of Bacillus subtilis that allows tRNA^Ile mischarging while retaining wild-type Ile-tRNA^Ile synthesis activity. As seen in other species defective for aminoacylation quality control, the growth rate of the ileS(T233P) strain was not significantly different from wild-type. When the ileS(T233P) strain was assessed for its ability to promote …

Ms-Read: Quantitative Measurement Of Amino Acid Incorporation, Kyle Mohler, Hans-Rudolph Aerni, Brandon Gassaway, Jiqiang Ling, Michael Ibba, Jesse Rinehart

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosomal protein synthesis results in the genetically programmed incorporation of amino acids into a growing polypeptide chain. Faithful amino acid incorporation that accurately reflects the genetic code is critical to the structure and function of proteins as well as overall proteome integrity. Errors in protein synthesis are generally detrimental to cellular processes yet emerging evidence suggest that proteome diversity generated through mistranslation may be beneficial under certain conditions. Cumulative translational error rates have been determined at the organismal level, however codon specific error rates and the spectrum of misincorporation errors from system to system remain largely unexplored. In particular, until …

Alpha-Synuclein: Insight Into The Hallmark Of Parkinson's Disease As A Target For Quantitative Molecular Diagnostics And Therapeutics, Baggio A. Evangelista

Honors Undergraduate Theses

Parkinson’s disease (PD) is the second-most common neurodegenerative disease after Alzheimer’s disease. With 500,000 individuals currently living with Parkinson’s and nearly 60,000 new cases diagnosed each year, this disease causes significant financial burden on the healthcare system - amassing to annual expenditures totaling 200 billion dollars; predicted to increase through 2050. The disease phenotype is characterized by a combination of a resting tremor, bradykinesia, muscular rigidity, and depression due to dopaminergic neuronal death in the midbrain. The cause of the neurotoxicity has been largely discussed, with strong evidence suggesting that the protein, alpha-Synuclein, is a key factor. Under native conditions, …

Micro-Spectroscopy Of Bio-Assemblies At The Single Cell Level, Jeslin Kera

Honors Undergraduate Theses

In this thesis, we investigate biological molecules on a micron scale in the ultraviolet spectral region through the non-destructive confocal absorption microscopy. The setup involves a combination of confocal microscope with a UV light excitation beam to measure the optical absorption spectra with spatial resolution of 1.4 μm in the lateral and 3.6 μm in the axial direction. Confocal absorption microscopy has the benefits of requiring no labels and only low light intensity for excitation while providing a strong signal from the contrast generated by the attenuation of propagating light due to absorption. This enables spatially resolved measurements of single …

Synthesis Of Rhamnosylated Arginine Glycopeptides And Determination Of The Glycosidic Linkage In Bacterial Elongation Factor P, Siyao Wang, Leo Corcilius, Phillip B. Sharp, Andrei Rajkovic, Michael Ibba, Benjamin L. Parker, Richard J. Payne

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

A new class of N-linked protein glycosylation – arginine rhamnosylation – has recently been discovered as a critical modification for the function of bacterial elongation factor P (EF-P). Herein, we describe the synthesis of suitably protected α- and β-rhamnosylated arginine amino acid “cassettes” that can be directly installed into rhamnosylated peptides. Preparation of a proteolytic fragment of Pseudomonas aeruginosa EF-P bearing both α- and β-rhamnosylated arginine enabled the unequivocal determination of the native glycosidic linkage to be α through 2D NMR and nano-UHPLC-tandem mass spectrometry studies.

Expression Of Glycine-Rich Proteins Found In Salivary Glands Of The Lone Star Tick (Amblyomma Americanum) Using A Mammalian Cell Line, Annabelle Clark

Honors Theses

Ticks play an important ecological role as well as a growing role in human health and veterinary care. Ticks are hosts to a plethora of microbial pathogens that can be transferred during feeding to cause tick-borne diseases in humans and many animals. Ticks may in large part owe the success of the transfer of these pathogens between hosts to their complex saliva. The saliva secreted upon a tick’s attachment to a host serves the following, among other, functions: anti-hemostasis of the blood pool, preventing an inflammatory response at the bite site, and serving as a natural anti-microbial substance. An important …

The Complex Evolutionary History Of Aminoacyl-Trna Synthetases, Anargyros Chaliotis, Panayotis Vlastaridis, Dimitris Mossialos, Michael Ibba, Hubert D. Becker, Constantinos Stathopoulos, Grigorios D. Amoutzias

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases (AARSs) are a superfamily of enzymes responsible for the faithful translation of the genetic code and have lately become a prominent target for synthetic biologists. Our large-scale analysis of >2500 prokaryotic genomes reveals the complex evolutionary history of these enzymes and their paralogs, in which horizontal gene transfer played an important role. These results show that a widespread belief in the evolutionary stability of this superfamily is misconceived. Although AlaRS, GlyRS, LeuRS, IleRS, ValRS are the most stable members of the family, GluRS, LysRS and CysRS often have paralogs, whereas AsnRS, GlnRS, PylRS and SepRS are often absent …

Isoacceptor Specific Characterization Of Trna Aminoacylation And Misacylation In Vivo, Kyle Mohler, Rebecca Mann, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Amino acid misincorporation during protein synthesis occurs due to misacylation of tRNAs or defects in decoding at the ribosome. While misincorporation of amino acids has been observed in a variety of contexts, less work has been done to directly assess the extent to which specific tRNAs are misacylated in vivo, and the identity of the misacylated amino acid moiety. Here we describe tRNA isoacceptor specific aminoacylation profiling (ISAP), a method to identify and quantify the amino acids attached to a tRNA species in vivo. ISAP allows compilation of aminoacylation profiles for specific isoacceptors tRNAs. To demonstrate the efficacy and …

Maintenance Of Transcription-Translation Coupling By Elongation Factor P, Sara Elgamal, Irina Artsimovitch, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Under conditions of tight coupling between translation and transcription, the ribosome enables synthesis of full-length mRNAs by preventing both formation of intrinsic terminator hairpins and loading of the transcription termination factor Rho. While previous studies have focused on transcription factors, we investigated the role of Escherichia coli elongation factor P (EF-P), an elongation factor required for efficient translation of mRNAs containing consecutive proline codons, in maintaining coupled translation and transcription. In the absence of EF-P, the presence of Rho utilization (rut) sites led to an ~30-fold decrease in translation of polyproline-encoding mRNAs. Coexpression of the Rho inhibitor Psu …

Translation Control Of Swarming Proficiency In Bacillus Subtilis By 5-Amino-Pentanolylated Elongation Factor P, Andrei Rajkovic, Katherine R. Hummels, Anne Witzky, Sarah Erickson, Philip R. Gafken, Julian P. Whitelegge, Kym F. Faull, Daniel B. Kearns, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) accelerates diprolyl synthesis and requires a posttranslational modification to maintain proteostasis. Two phylogenetically distinct EF-P modification pathways have been described and are encoded in the majority of Gram-negative bacteria, but neither is present in Gram-positive bacteria. Prior work suggested that the EF-P-encoding gene (efp) primarily supports Bacillus subtilis swarming differentiation, whereas EF-P in Gram-negative bacteria has a more global housekeeping role, prompting our investigation to determine whether EF-P is modified and how it impacts gene expression in motile cells. We identified a 5-aminopentanol moiety attached to Lys³² of B. subtilis EF-P that is …

Multiple Quality Control Pathways Limit Non-Protein Amino Acid Use By Yeast Cytoplasmic Phenylalanyl-Trna Synthetase, Adil Moghal, Lin Hwang, Kym F. Faull, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Non-protein amino acids, particularly isomers of the proteinogenic amino acids, present a threat to proteome integrity if they are mistakenly inserted into proteins. Quality control during aminoacyl-tRNA synthesis reduces non-protein amino acid incorporation by both substrate discrimination and proofreading. For example phenylalanyl-tRNA synthetase (PheRS) proofreads the non-protein hydroxylated phenylalanine derivative m-Tyr after its attachment to tRNA^Phe. We now show in Saccharomyces cerevisiae that PheRS misacylation of tRNA^Phe with the more abundant Phe oxidation product o-Tyr is limited by kinetic discrimination against o-Tyr-AMP in the transfer step followed by o-Tyr-AMP release from the synthetic …

Non-Canonical Roles Of Trnas And Trna Mimics In Bacterial Cell Biology, Assaf Katz, Sara Elgamal, Andrei Rajkovic, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Transfer RNAs (tRNAs) are the macromolecules that transfer activated amino acids from aminoacyl‐tRNA synthetases to the ribosome, where they are used for the mRNA guided synthesis of proteins. Transfer RNAs are ancient molecules, perhaps even predating the existence of the translation machinery. Albeit old, these molecules are tremendously conserved, a characteristic that is well illustrated by the fact that some bacterial tRNAs are efficient and specific substrates of eukaryotic aminoacyl‐tRNA synthetases and ribosomes. Considering their ancient origin and high structural conservation, it is not surprising that tRNAs have been hijacked during evolution for functions outside of translation. These roles beyond …

Modulation Of Cell Death Signaling And Cell Proliferation By The Interaction Of Homoserine Lactones And Paraoxonase 2., Aaron Mackallan Neely

Electronic Theses and Dissertations

Pseudomonas aeruginosa produces N-(3-oxododecanoyl)-homoserine lactone (C12) as a quorum-sensing molecule that functions to facilitate bacteria-bacteria communication. C12 has also been reported to affect many aspects of human host cell physiology, including evoking cell death in various types of cells. However, the signaling pathway(s) leading to C12-triggerred cell death remains unclear. To clarify cell death signaling induced by C12, we examined mouse embryonic fibroblasts (MEFs) deficient in one or more caspases. Our data indicate that, unlike most apoptotic inducers, C12 evokes a novel form of apoptosis in cells, probably through the direct induction of mitochondrial membrane permeabilization. Previous studies indicate that …

Engineering A Mutation In The Heparin Binding Pocket Of The Human Fibroblast Growth Factor, Roshni Patel

Chemistry & Biochemistry Undergraduate Honors Theses

Fibroblast growth factors (FGFs) are family of proteins that belong to a group of growth factors that are found in mammals and play an important role in angiogenesis, differentiation, organogenesis, and tissue repair. In summary, their main functionality is involved in cell division and proliferation. Because FGFs plays such a vital role in cell proliferation, they are mainly involved in the process of wound healing and injuries. FGF binds to its ligand, heparin—a heavily sulfated glycosaminoglycan. The binding of heparin to FGF occurs through electrostatic interactions, specifically between the negatively charged sulfate groups on heparin and positively charged residues such …

Novel Compound Heterozygous Mutations Expand The Recognized Phenotypes Of Fars2-Linked Disease, Melissa A. Walker, Kyle Mohler, Kyle W. Hopkins, Derek H. Oakley, David A. Sweetser, Michael Ibba, Matthew P. Frosch, Ronald L. Thibert

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Mutations in mitochondrial aminoacyl-tRNA synthetases are an increasingly recognized cause of human diseases, often arising in individuals with compound heterozygous mutations and presenting with system-specific phenotypes, frequently neurologic. FARS2 encodes mitochondrial phenylalanyl transfer ribonucleic acid (RNA) synthetase (mtPheRS), perturbations of which have been reported in 6 cases of an infantile, lethal disease with refractory epilepsy and progressive myoclonus. Here the authors report the case of juvenile onset refractory epilepsy and progressive myoclonus with compound heterozygous FARS2 mutations. The authors describe the clinical course over 6 years of care at their institution and diagnostic studies including electroencephalogram (EEG), brain magnetic resonance …

Confirmation Of Emx2 Protein Binding Partners, Cody Gillman

All Master's Theses

The neocortex is a structure within mammalian brains that processes sensory input from eyes, ears, and touch receptors and mediates the conscious use of skeletal muscles. The processing of information related to each of these types of functions is localized within discrete areas of the neocortex, which are separated by sharp borders. Proper development of these functional areas is regulated during embryogenesis by several transcription factors that are expressed in distinct gradients across the progenitor layer of the neocortex, the ventricular zone. Despite the vast amount of progress that has been made in describing how these transcription factors impact the …

Enabling Method To Design Versatile Biomaterial Systems From Colloidal Building Blocks, Shalini Saxena, L. Andrew Lyon

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Development of materials with fine spatial control over topographical, mechanical, or chemical features has been investigated for a variety of applications. Here we present a method to fabricate an array of polyelectrolyte constructs including two-dimensionally and three-dimensionally patterned assemblies using both compressible and incompressible colloidal building blocks. This method eliminates prior constraints associated with specific chemistries, and can be used to develop modular, multi-component, patterned assemblies. In particular, development of constructs were investigated using microgels, which are colloidally stable hydrogel microparticles, polystyrene (PS) beads, and PS-microgel core-shell building blocks in conjunction with the polycation poly(ethyleneimine) (PEI). The topography, mechanical properties, …

Inactivation Of Cytomegalovirus In Breast Milk Using Ultraviolet-C Irradiation: Opportunities For A New Treatment Option In Breast Milk Banking, Megan L. Lloyd, Nurul Hod, Jothsna Jayaraman, Elizabeth A. Marchant, Lukas Christen, Peter Chiang, Peter E. Hartmann, Geoffrey R. Shellam, Karen N. Simmer

Research outputs 2014 to 2021

Pasteurized donor human milk is provided by milk banks to very preterm babies where their maternal supply is insufficient or unavailable. Donor milk is currently processed by Holder pasteurization, producing a microbiologically safe product but significantly reducing immunoprotective components. Ultraviolet-C (UV-C) irradiation at 254 nm is being investigated as an alternative treatment method and has been shown to preserve components such as lactoferrin, lysozyme and secretory IgA considerably better than Holder pasteurization. We describe the inactivation of cytomegalovirus, a virus commonly excreted into breast milk, using UV-C irradiation. Full replication was ablated by various treatment doses. However, evidence of viral …

Elongation Factor-P At The Crossroads Of The Host-Endosymbiont Interface, Andrei Rajkovic, Anne Witzky, William Navarre, Andrew J. Darwin, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is an ancient bacterial translational factor that aids the ribosome in polymerizing oligo-prolines. EF-P structurally resembles tRNA and binds in-between the exit and peptidyl sites of the ribosome to accelerate the intrinsically slow reaction of peptidyl-prolyl bond formation. Recent studies have identified in separate organisms, two evolutionarily convergent EF-P post-translational modification systems (EPMS), split predominantly between gammaproteobacteria, and betaproteobacteria. In both cases EF-P receives a post-translational modification, critical for its function, on a highly conserved residue that protrudes into the peptidyl-transfer center of the ribosome. EPMSs are comprised of a gene(s) that synthesizes the precursor molecule …

Elucidating The Role Of Hausp Ubiquitin Like Domains In The Catalytic Function Of Usp7, Anuj Patel, Nicole Davis, Andrew Mesecar

The Summer Undergraduate Research Fellowship (SURF) Symposium

Ubiquitin specific proteases (USPs) are a class of enzymes involved in myriad cellular processes. One USP of great interest due to its oncogenic properties is USP7. In normal conditions USP7 is closely regulated due to its responsibility for destabilizing the tumor suppressor, p53, through the deubiquitination of MDM2. In multiple myeloma cases, it appears the regulation of USP7 subsides, as it is largely overexpressed, leading to the inappropriate degradation of p53. Inhibition of USP7 could, therefore, prove a viable target for cancer therapy. A greater understanding of USP7’s function and structure can lead to more insight into how this enzyme …

Cyclic Rhamnosylated Elongation Factor P Establishes Antibiotic Resistance In Pseudomonas Aeruginosa, Andrei Rajkovic, Sarah Erickson, Anne Witzky, Owen E. Branson, Jin Seo, Philip R. Gafken, Michael A. Frietas, Julian P. Whitelegge, Kym F. Faull, William Wiley Navarre, Andrew J. Darwin, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is a ubiquitous bacterial protein that is required for the synthesis of poly-proline motifs during translation. In Escherichia coli and Salmonella enterica, the posttranslational β-lysylation of Lys34 by the PoxA protein is critical for EF-P activity. PoxA is absent from many bacterial species such as Pseudomonas aeruginosa, prompting a search for alternative EF-P posttranslation modification pathways. Structural analyses of P. aeruginosa EF-P revealed the attachment of a single cyclic rhamnose moiety to an Arg residue at a position equivalent to that at which β-Lys is attached to E. coli EF-P. Analysis of the genomes …

Transfer Rna Comes Of Age, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

"The year the journal RNA was founded was slated by some in scientific publishing to be the year that one particular type of RNA's run in the spotlight would end. In 1995 I had recently started as a post-doc with Dieter Söll at Yale when he came into the lab to solemnly inform us all that an editor at a certain (S)cience journal had just told him “we won't be publishing any more tRNA papers.” For a post-doc who had migrated across the Atlantic for the sole purpose of furthering his career by working on tRNA this was not great …

Effect Of Hydrogen Peroxide On The Biosynthesis Of Heme And Proteins: Potential Implications For The Partitioning Of Glu-TrnaGlu Between These Pathways, Carolina Farah, Gloria Levicán, Michael Ibba, Omar Orellana

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Glutamyl-tRNA (Glu-tRNA^Glu) is the common substrate for both protein translation and heme biosynthesis via the C5 pathway. Under normal conditions, an adequate supply of this aminoacyl-tRNA is available to both pathways. However, under certain circumstances, Glu-tRNA^Glu can become scarce, resulting in competition between the two pathways for this aminoacyl-tRNA. In Acidithiobacillus ferrooxidans, glutamyl-tRNA synthetase 1 (GluRS1) is the main enzyme that synthesizes Glu-tRNA^Glu. Previous studies have shown that GluRS1 is inactivated in vitro by hydrogen peroxide (H₂O₂). This raises the question as to whether H₂O₂ negatively affects …

Numerical Simulations Of In Vitro Nanoparticle Toxicity – The Case Of Poly(Amido Amine) Dendrimers., Marcus Maher, Pratap Naha, Sourav Prasanna Mukherjee, Hugh Byrne

Articles

A phenomenological rate equation model is constructed to numerically simulate nanoparticle uptake and subsequent cellular response. Polyamidoamine dendrimers (generations 4-6) are modelled and the temporal evolution of the intracellular cascade of; increased levels of reactive oxygen species, intracellular antioxidant species, caspase activation, mitochondrial membrane potential decay, tumour necrosis factor and interleukin generation is simulated, based on experimental observations.

The dose and generation dependence of several of these response factors are seen to well represent experimental observations at a range of time points. The model indicates that variations between responses of different cell-lines, including murine macrophages, human keratinocytes and colon cells, …

The Non-Canonical Hydroxylase Structure Of Yfcm Reveals A Metal Ion-Coordination Motif Required For Ef-P Hydroxylation, Kan Kobayashi, Assaf Katz, Andrei Rajkovic, Ryohei Ishii, Owen E. Branson, Michael A. Freitas, Ryuichiro Ishitani, Michael Ibba, Osamu Nureki

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

EF-P is a bacterial tRNA-mimic protein, which accelerates the ribosome-catalyzed polymerization of poly-prolines. In Escherichia coli, EF-P is post-translationally modified on a conserved lysine residue. The post-translational modification is performed in a two-step reaction involving the addition of a β-lysine moiety and the subsequent hydroxylation, catalyzed by PoxA and YfcM, respectively. The β-lysine moiety was previously shown to enhance the rate of poly-proline synthesis, but the role of the hydroxylation is poorly understood. We solved the crystal structure of YfcM and performed functional analyses to determine the hydroxylation mechanism. In addition, YfcM appears to be structurally distinct from any …

Mistranslation Of The Genetic Code, Adil Moghal, Kyle Mohler, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

During mRNA decoding at the ribosome, deviations from stringent codon identity, or “mistranslation,” are generally deleterious and infrequent. Observations of organisms that decode some codons ambiguously, and the discovery of a compensatory increase in mistranslation frequency to combat environmental stress have changed the way we view “errors” in decoding. Modern tools for the study of the frequency and phenotypic effects of mistranslation can provide quantitative and sensitive measurements of decoding errors that were previously inaccessible. Mistranslation with non‐protein amino acids, in particular, is an enticing prospect for new drug therapies and the study of molecular evolution.

Relaxed Substrate Specificity Leads To Extensive Trna Mischarging By Streptococcus Pneumoniae Class I And Class Ii Aminoacyl-Trna Synthetases, Jennifer Shepherd, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases provide the first step in protein synthesis quality control by discriminating cognate from noncognate amino acid and tRNA substrates. While substrate specificity is enhanced in many instances by cis- and trans-editing pathways, it has been revealed that in organisms such as Streptococcus pneumoniae some aminoacyl-tRNA synthetases display significant tRNA mischarging activity. To investigate the extent of tRNA mischarging in this pathogen, the aminoacylation profiles of class I isoleucyl-tRNA synthetase (IleRS) and class II lysyl-tRNA synthetase (LysRS) were determined. Pneumococcal IleRS mischarged tRNA^Ile with both Val, as demonstrated in other bacteria, and Leu in a tRNA sequence-dependent …

Translation Initiation Rate Determines The Impact Of Ribosome Stalling On Bacterial Protein Synthesis, Steven J. Hersch, Sara Elgamal, Assaf Katz, Michael Ibba, William Wiley Navarre

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosome stalling during translation can be caused by a number of characterized mechanisms. However, the impact of elongation stalls on protein levels is variable, and the reasons for this are often unclear. To investigate this relationship, we examined the bacterial translation elongation factor P (EF-P), which plays a critical role in rescuing ribosomes stalled at specific amino acid sequences including polyproline motifs. In previous proteomic analyses of both Salmonella and Escherichia coli efp mutants, it was evident that not all proteins containing a polyproline motif were dependent on EF-P for efficient expression in vivo . The α- and β-subunits of …