Set Of Codons Research Articles

Codon usage bias of human papillomavirus type 33 and 58: A comprehensive analysis.

Cervical cancer is closely linked to specific strains of human papillomavirus (HPV), notably HPV-33 and HPV-58, which exhibit a significant prevalence among women in China. Nevertheless, the codon usage bias in HPV-33 and HPV-58 is not well comprehended. The objective of this research is to analyze the codon usage patterns HPV-33 and HPV-58, pinpoint the primary factors that influence codon preference. The overall preference for codon usage in two HPV genotypes is not significant. Both HPV genotypes exhibit a preference for codons that end with A/U. The GC3 content for HPV-33 is 25.43% ± 0.35%, and for HPV-58, it is 29.44% ± 0.57%. Out of the 26 favored codons in HPV-33 and HPV-58 (relative synonymous codon usage (RSCU) > 1), 25 conclude with A/U. Principal component analysis (PCA) shows a tight clustering of the entire genome sequences of HPV-33 and HPV-58, suggesting a similarity in their RSCU preferences. Moreover, an examination of dinucleotide abundance indicated that translation selection influenced the development of a distinctive dinucleotide usage pattern in HPV-33 and HPV-58. Additionally, a combined analysis involving an effective number of codons plot, parity rule 2, and neutrality analysis demonstrated that, for HPV-33 and HPV-58, the primary determinant influencing codon usage preference is natural selection. HPV-33 and HPV-58 exhibit a restricted set of favored codons in common with humans, potentially mitigating competition for translation resources. Our discoveries could provide valuable perspectives on the evolutionary patterns and codon usage preferences of HPV-33 and HPV-58 viruses, contributing to the development and application of relevant HPV subtype vaccines.

Significance of genetic code module structure in gene expression and GC content enhancement in RNA sequences

The existent algebraic models of the genetic code contribute to the understanding of the physio-chemical characteristics of the amino acids. However, the process of translating a gene into a phenotype is highly complex. Moreover, the intricacy of gene expression gets further multiplied due to the biases in the codon usage. This paper explores an algebraic structure called module on the set of codons as well as on that of RNA sequences. We study the potential implications of these structures on gene expression and the GC content of an RNA sequence. The base order {C,U,G,A} appears to possess greater biological significance than many of the orders previously studied. We have developed a novel algorithm to generate RNA sequences with high GC content, aiming to enhance the thermostability of biomolecules. The insights gained from this investigation may have applications in biomolecular modeling and docking, protein engineering, drug development, and related fields.

Statistical analysis of synonymous and stop codons in pseudo-random and real sequences as a function of GC content

Knowledge of the frequencies of synonymous triplets in protein-coding and non-coding DNA stretches can be used in gene finding. These frequencies depend on the GC content of the genome or parts of it. An example of interest is provided by stop codons. This is relevant for the definition of Open Reading Frames. A generic case is provided by pseudo-random sequences, especially when they code for complex proteins or when they are non-coding and not subject to selection pressure. Here, we calculate, for such sequences and for all 25 known genetic codes, the frequency of each amino acid and stop codon based on their set of codons and as a function of GC content. The amino acids can be classified into five groups according to the GC content where their expected frequency reaches its maximum. We determine the overall Shannon information based on groups of synonymous codons and show that it becomes maximum at a percent GC of 43.3% (for the standard code). This is in line with the observation that in most fungi, plants, and animals, this genomic parameter is in the range from 35 to 50%. By analysing natural sequences, we show that there is a clear bias for triplets corresponding to stop codons near the 5′- and 3′-splice sites in the introns of various clades.

Growth-dependent Gene Expression Variation Influences the Strength of Codon Usage Biases.

The most highly expressed genes in microbial genomes tend to use a limited set of synonymous codons, often referred to as "preferred codons." The existence of preferred codons is commonly attributed to selection pressures on various aspects of protein translation including accuracy and/or speed. However, gene expression is condition-dependent and even within single-celled organisms transcript and protein abundances can vary depending on a variety of environmental and other factors. Here, we show that growth rate-dependent expression variation is an important constraint that significantly influences the evolution of gene sequences. Using large-scale transcriptomic and proteomic data sets in Escherichia coli and Saccharomyces cerevisiae, we confirm that codon usage biases are strongly associated with gene expression but highlight that this relationship is most pronounced when gene expression measurements are taken during rapid growth conditions. Specifically, genes whose relative expression increases during periods of rapid growth have stronger codon usage biases than comparably expressed genes whose expression decreases during rapid growth conditions. These findings highlight that gene expression measured in any particular condition tells only part of the story regarding the forces shaping the evolution of microbial gene sequences. More generally, our results imply that microbial physiology during rapid growth is critical for explaining long-term translational constraints.

Decrypting pMHCI Diversity Through Noncanonical Translation Initiation

Abstract CD8 +T-cells rely on MHCI to surveil the proteomic state of cells in the form of peptide-MHCI (pMHCI). We previously uncovered a set of ATG-like translation initiation codons that can define noncanonical open reading frames (ORFs) of cryptic peptides presented by MHCI. The model cryptic codon CTG shares key translation initiation requirements with canonical ATG initiation, yet incorporates N-terminal Methionine (nMet) or Leucine (nLeu). We aim to characterize the activity and peptide products of the remaining cryptic codons to understand how cryptic ORFs contribute to the peptidome accessible to MHCI. Therefore, we make use of the subfemtomole sensitivity of the hyperB3Z reporter T-cell hybridoma to interrogate the identity of a modified OVA peptide (SIINFEHL;[SHL8]) reporter in a fixed cryptic initiation context, while retaining information on the residue incorporated during translation initiation. Following transfection of mammalian model cell lines with reporter, purified peptides from total cell extracts are RP-HPLC fractionated to differentiate N-terminally extended and trimmed [SHL8] intermediates. We found that nMet is incorporated by CTG and ACG initiation, while nLeu was absent under CTG initiation despite coexpression of the H2-D ballotype known to protect N-terminally extended [SHL8]. Peptide reporters with a P-2 residue derived from a codon satisfying the optimal (CACCxxx G) Kozak sequence (i.e. G, D) revealed that bulky P-2 residues enhance recovery of total reporter suggesting cryptic ORF peptide stability largely depends on P-2. Thus, by understanding the products of cryptic ORFs we expand the definition of immune-self while exposing new sources of pMHCI unique to autoimmunity and viral pathogens. Supported by grants from NIH (R01-AI130210, R01-AI121174, R37-AI060040)

Perseverance of protein homeostasis despite mistranslation of glycine codons with alanine.

By linking amino acids to their codon assignments, transfer RNAs (tRNAs) are essential for protein synthesis and translation fidelity. Some human tRNA variants cause amino acid mis-incorporation at a codon or set of codons. We recently found that a naturally occurring tRNASer variant decodes phenylalanine codons with serine and inhibits protein synthesis. Here, we hypothesized that human tRNA variants that misread glycine (Gly) codons with alanine (Ala) will also disrupt protein homeostasis. The A3G mutation occurs naturally in tRNAGly variants (tRNAGlyCCC, tRNAGlyGCC) and creates an alanyl-tRNA synthetase (AlaRS) identity element (G3 : U70). Because AlaRS does not recognize the anticodon, the human tRNAAlaAGC G35C (tRNAAlaACC) variant may function similarly to mis-incorporate Ala at Gly codons. The tRNAGly and tRNAAla variants had no effect on protein synthesis in mammalian cells under normal growth conditions; however, tRNAGlyGCC A3G depressed protein synthesis in the context of proteasome inhibition. Mass spectrometry confirmed Ala mistranslation at multiple Gly codons caused by the tRNAGlyGCC A3G and tRNAAlaAGC G35C mutants, and in some cases, we observed multiple mistranslation events in the same peptide. The data reveal mistranslation of Ala at Gly codons and defects in protein homeostasis generated by natural human tRNA variants that are tolerated under normal conditions. This article is part of the theme issue 'Reactivity and mechanism in chemical and synthetic biology'.

A novel fluorescent reporter sensitive to serine mis-incorporation

ABSTRACT High-fidelity translation was considered a requirement for living cells. The frozen accident theory suggested that any deviation from the standard genetic code should result in the production of so much mis-made and non-functional proteins that cells cannot remain viable. Studies in bacterial, yeast, and mammalian cells show that significant levels of mistranslation (1–10% per codon) can be tolerated or even beneficial under conditions of oxidative stress. Single tRNA mutants, which occur naturally in the human population, can lead to amino acid mis-incorporation at a codon or set of codons. The rate or level of mistranslation can be difficult or impossible to measure in live cells. We developed a novel red fluorescent protein reporter that is sensitive to serine (Ser) mis-incorporation at proline (Pro) codons. The mCherry Ser151Pro mutant is efficiently produced in Escherichia coli but non-fluorescent. We demonstrated in cells and with purified mCherry protein that the fluorescence of mCherry Ser151Pro is rescued by two different tRNASer gene variants that were mutated to contain the Pro (UGG) anticodon. Ser mis-incorporation was confirmed by mass spectrometry. Remarkably, E. coli tolerated mistranslation rates of ~10% per codon with negligible reduction in growth rate. Conformational sampling simulations revealed that the Ser151Pro mutant leads to significant changes in the conformational freedom of the chromophore precursor, which is indicative of a defect in chromophore maturation. Together our data suggest that the mCherry Ser151 mutants may be used to report Ser mis-incorporation at multiple other codons, further expanding the ability to measure mistranslation in living cells.

Highly efficient libraries design for saturation mutagenesis.

Saturation mutagenesis is a semi-rational approach for protein engineering where sites are saturated either entirely or partially to include amino acids of interest. We previously reported on a codon compression algorithm, where a set of minimal degenerate codons are selected according to user-defined parameters such as the target organism, type of saturation and usage levels. Here, we communicate an addition to our web tool that considers the distance between the wild-type codon and the library, depending on its purpose. These forms of restricted collections further reduce library size, lowering downstream screening efforts or, in turn, allowing more comprehensive saturation of multiple sites. The library design tool can be accessed via http://www.dynamcc.com/dynamcc_d/. Graphical

Codes over Lattice‐Valued Intuitionistic Fuzzy Set Type‐3 with Application to the Complex DNA Analysis

In this article codes over lattice valued intuitionistic fuzzy set type‐3 (LIFS‐3) are defined. Binary block codes and linear codes are constructed over LIFS‐3. Hamming distance and related properties of these newly established codes are examined. The research findings are applied to genetic codes. The set L of sixty‐four codons is converted into a lattice and then codes are created over the set S of twenty amino acids by defining membership and nonmembership functions from the set of twenty amino acids to the sixty‐four codon set. Comparison of codes over L‐fuzzy set and LIFS‐3 conducted in terms of hamming distance for codon system that ensures the efficiency of newly established codes.

Codon Usage of Chloroplast Gene rbcL in Laurencia sensu lato (Rhodophyta) species

Recent advances in molecular biology make it possible to sequence not only genes or genomes, but also to understand codon dynamics. For the organelle genes of these organisms, a small set of preferred codons are used for encoding proteins. For the first time, this paper treats the divergence of synonymous codon usage and its bias in the rbcL gene within the Laurencia complex of red algae. We observed that the synonymous codon preference biases in rbcL are large and differ among species. A clear distinction in codon usage between genera is evident: the genera Dasya and Delesseria use a set that fluctuated between 53 and 58 codons. Whereas, in the genera Ceramium, Chondrophycus, Chlamydomonas, Chlorella, Laurencia s.s, Osmundea, and Palisada codon usage indicates a higher restriction fluctuating between 40 to 51 codons. Laurencia complex genera and other representative algae showed a defined composition pattern, with lower percentage values of NNC/G (7-24.9%). Dasya and Delesseria showed a selective pattern tendency because of high percentage values of NNC/G (54-55%). The estimated codon bias parameters were tested to infer systematic relationships and match suitable codons with the NNC / G codon percentages. Cluster analysis based on Codon Usage supports phylogenetic relationships between Chondrophycus, Palisada, Laurencia, Osmundea, and Yuzurua species.

The impact of codon choice on translation process in Saccharomyces cerevisiae: folding class, protein function and secondary structure

The genetic code consists in a set of rules used by living organisms to translate genomic information, contained in genes, into proteins; every amino acid is coded by a set of nucleotide triplets or codons. We refer to codon choice as the choice of a given codon, among the synonymous available ones, to code a given amino acid occurrence.The aim of this work is to shed light on the pivotal role that codon choice plays in regulating the timing of translation process, through patterns of low and high translation efficiency codons. A translation efficiency value, namely codon score, was associated to each codon through a formula based on the number of tRNAs gene copies able to translate the given codon. By using codon scores, those k-mers of the proteome of Saccharomyces cerevisiae, showing low and high average scores associated to the correspondent codons, were computed.The analysis of distribution of both low and high average score k-mers clearly showed that, in particular for higher k-mer size, they occur much more than expected, strongly suggesting a functional role. Moreover performed analysis highlighted that significant k-mers preferentially occur in some protein folding classes, such as those containing alpha helices, and in some functional classes mainly involved in transcription process while codon choice seems to have a very low impact in proteins associated to energy production and metabolism. The relationship between secondary structures and significant k-mers was investigated, revealing that low score k-mers tend to preferentially occur in coil or close to coil regions and almost never in beta sheets, while high score k-mers preferentially occur in alpha helices, avoiding beta sheets, and close to coil regions for high k-mer sizes.Finally the analysis of distribution of significant codon patterns along the proteins highlighted a relevant enrichment of low average score k-mers at the 5’ end of protein-coding sequences in the region from 5th to 25th amino acid.

Some theoretical aspects of reprogramming the standard genetic code.

Reprogramming of the standard genetic code to include non-canonical amino acids (ncAAs) opens new prospects for medicine, industry, and biotechnology. There are several methods of code engineering, which allow us for storing new genetic information in DNA sequences and producing proteins with new properties. Here, we provided a theoretical background for the optimal genetic code expansion, which may find application in the experimental design of the genetic code. We assumed that the expanded genetic code includes both canonical and non-canonical information stored in 64 classical codons. What is more, the new coding system is robust to point mutations and minimizes the possibility of reversion from the new to old information. In order to find such codes, we applied graph theory to analyze the properties of optimal codon sets. We presented the formal procedure in finding the optimal codes with various number of vacant codons that could be assigned to new amino acids. Finally, we discussed the optimal number of the newly incorporated ncAAs and also the optimal size of codon groups that can be assigned to ncAAs.

Coding triplets in the tRNA acceptor-TΨC arm and their role in present and past tRNA recognition.

The mechanism and evolution of the recognition scheme between key components of the translation system, that is, tRNAs, synthetases, and elongation factors, are fundamental issues in understanding the translation of genetic information into proteins. Statistical analysis of bacterial tRNA sequences reveals that for six amino acids, a string of 10 nucleotides preceding the tRNA 3' end carries cognate coding triplets to nearly full extent. The triplets conserved in positions 63-67 are implicated in the recognition by the elongation factor EF-Tu, and those conserved in positions 68-72, in the identification of cognate tRNAs, and their derived minihelices by class IIa synthetases. These coding triplets are suggested to have primordial origin, being engaged in aminoacylation of prebiotic tRNAs and in the establishment of the canonical codon set.

Codon-based indices for modeling gene expression and transcript evolution.

Codon usage bias (CUB) refers to the phenomena that synonymous codons are used in different frequencies in most genes and organisms. The general assumption is that codon biases reflect a balance between mutational biases and natural selection. Today we understand that the codon content is related and can affect all gene expression steps. Starting from the 1980s, codon-based indices have been used for answering different questions in all biomedical fields, including systems biology, agriculture, medicine, and biotechnology. In general, codon usage bias indices weigh each codon or a small set of codons to estimate the fitting of a certain coding sequence to a certain phenomenon (e.g., bias in codons, adaptation to the tRNA pool, frequencies of certain codons, transcription elongation speed, etc.) and are usually easy to implement. Today there are dozens of such indices; thus, this paper aims to review and compare the different codon usage bias indices, their applications, and advantages. In addition, we perform analysis that demonstrates that most indices tend to correlate even though they aim to capture different aspects. Due to the centrality of codon usage bias on different gene expression steps, it is important to keep developing new indices that can capture additional aspects that are not modeled with the current indices.

Arming Oncolytic Adenoviruses: Effect of Insertion Site and Splice Acceptor on Transgene Expression and Viral Fitness.

Oncolytic adenoviruses (OAds) present limited efficacy in clinics. The insertion of therapeutic transgenes into OAds genomes, known as “arming OAds”, has been the main strategy to improve their therapeutic potential. Different approaches were published in the decade of the 2000s, but with few comparisons. Most armed OAds have complete or partial E3 deletions, leading to a shorter half-life in vivo. We generated E3+ OAds using two insertion sites, After-fiber and After-E4, and two different splice acceptors linked to the major late promoter, either the Ad5 protein IIIa acceptor (IIIaSA) or the Ad40 long fiber acceptor (40SA). The highest transgene levels were obtained with the After-fiber location and 40SA. However, the set of codons of the transgene affected viral fitness, highlighting the relevance of transgene codon usage when arming OAds using the major late promoter.

Universal Codons with Enrichment from GC to AU Nucleotide Composition Reveal a Chronological Assignment from Early to Late Along with LUCA Formation.

The emergence of a primitive genetic code should be considered the most essential event during the origin of life. Almost a complete set of codons (as we know them) should have been established relatively early during the evolution of the last universal common ancestor (LUCA) from which all known organisms descended. Many hypotheses have been proposed to explain the driving forces and chronology of the evolution of the genetic code; however, none is commonly accepted. In the current paper, we explore the features of the genetic code that, in our view, reflect the mechanism and the chronological order of the origin of the genetic code. Our hypothesis postulates that the primordial RNA was mostly GC-rich, and this bias was reflected in the order of amino acid codon assignment. If we arrange the codons and their corresponding amino acids from GC-rich to AU-rich, we find that: 1. The amino acids encoded by GC-rich codons (Ala, Gly, Arg, and Pro) are those that contribute the most to the interactions with RNA (if incorporated into short peptides). 2. This order correlates with the addition of novel functions necessary for the evolution from simple to longer folded peptides. 3. The overlay of aminoacyl-tRNA synthetases (aaRS) to the amino acid order produces a distinctive zonal distribution for class I and class II suggesting an interdependent origin. These correlations could be explained by the active role of the bridge peptide (BP), which we proposed earlier in the evolution of the genetic code.

Basic principles of the genetic code extension.

Compounds including non-canonical amino acids (ncAAs) or other artificially designed molecules can find a lot of applications in medicine, industry and biotechnology. They can be produced thanks to the modification or extension of the standard genetic code (SGC). Such peptides or proteins including the ncAAs can be constantly delivered in a stable way by organisms with the customized genetic code. Among several methods of engineering the code, using non-canonical base pairs is especially promising, because it enables generating many new codons, which can be used to encode any new amino acid. Since even one pair of new bases can extend the SGC up to 216 codons generated by a six-letter nucleotide alphabet, the extension of the SGC can be achieved in many ways. Here, we proposed a stepwise procedure of the SGC extension with one pair of non-canonical bases to minimize the consequences of point mutations. We reported relationships between codons in the framework of graph theory. All 216 codons were represented as nodes of the graph, whereas its edges were induced by all possible single nucleotide mutations occurring between codons. Therefore, every set of canonical and newly added codons induces a specific subgraph. We characterized the properties of the induced subgraphs generated by selected sets of codons. Thanks to that, we were able to describe a procedure for incremental addition of the set of meaningful codons up to the full coding system consisting of three pairs of bases. The procedure of gradual extension of the SGC makes the whole system robust to changing genetic information due to mutations and is compatible with the views assuming that codons and amino acids were added successively to the primordial SGC, which evolved minimizing harmful consequences of mutations or mistranslations of encoded proteins.

EIF1 discriminates against suboptimal initiation sites to prevent excessive uORF translation genome-wide.

The translation preinitiation complex (PIC) scans the mRNA for an AUG codon in a favorable context. Previous findings suggest that the factor eIF1 discriminates against non-AUG start codons by impeding full accommodation of Met-tRNAi in the P site of the 40S ribosomal subunit, necessitating eIF1 dissociation for start codon selection. Consistent with this, yeast eIF1 substitutions that weaken its binding to the PIC increase initiation at UUG codons on a mutant his4 mRNA and particular synthetic mRNA reporters; and also at the AUG start codon of the mRNA for eIF1 itself owing to its poor Kozak context. It was not known however whether such eIF1 mutants increase initiation at suboptimal start codons genome-wide. By ribosome profiling, we show that the eIF1-L96P variant confers increased translation of numerous upstream open reading frames (uORFs) initiating with either near-cognate codons (NCCs) or AUGs in poor context. The increased uORF translation is frequently associated with the reduced translation of the downstream main coding sequences (CDS). Initiation is also elevated at certain NCCs initiating amino-terminal extensions, including those that direct mitochondrial localization of the GRS1 and ALA1 products, and at a small set of main CDS AUG codons with especially poor context, including that of eIF1 itself. Thus, eIF1 acts throughout the yeast translatome to discriminate against NCC start codons and AUGs in poor context; and impairing this function enhances the repressive effects of uORFs on CDS translation and alters the ratios of protein isoforms translated from near-cognate versus AUG start codons.

Comparative in silico analysis of ftsZ gene from different bacteria reveals the preference for core set of codons in coding sequence structuring and secondary structural elements determination.

The deluge of sequence information in the recent times provide us with an excellent opportunity to compare organisms on a large genomic scale. In this study we have tried to decipher the variation in the gene organization and structuring of a vital bacterial gene called ftsZ which codes for an integral component of the bacterial cell division, the FtsZ protein. FtsZ is homologous to tubulin protein and has been found to be ubiquitous in eubacteria. FtsZ is showing increasing promise as a target for antibacterial drug discovery. Our study of ftsZ protein from 143 different bacterial species spanning a wider range of morphological and physiological type demonstrates that the ftsZ gene of about ninety three percent of the organisms show relatively biased codon usage profile and significant GC deviation from their genomic GC content. Comparative codon usage analysis of ftsZ and a core housekeeping gene rpoB demonstrated that codon usage pattern of ftsZ CDS is shaped by natural selection to a large extent and mimics that of a housekeeping gene. We have also detected a tendency among the different organisms to utilize a core set of codons in structuring the ftsZ coding sequence. We observed that the compositional frequency of the amino acid serine in the FtsZ protein appears to be a indicator of the bacterial lifestyle. Our meticulous analysis of the ftsZ gene linked with the corresponding FtsZ protein show that there is a bias towards the use of specific synonymous codons particularly in the helix and strand regions of the multi-domain FtsZ protein. Overall our findings suggest that in an indispensable and vital protein such as FtsZ, there is an inherent tendency to maintain form for optimized performance in spite of the extrinsic variability in coding features.

Serine substitutions are linked to codon usage and differ for variable and conserved protein regions

Serine is the only amino acid that is encoded by two disjoint codon sets (TCN & AGY) so that a tandem substitution of two nucleotides is required to switch between the two sets. We show that these codon sets underlie distinct substitution patterns at positions subject to purifying and diversifying selections. We found that in humans, positions that are conserved among ~100 vertebrates, and thus subjected to purifying selection, are enriched for substitutions involving serine (TCN, denoted S′), proline, and alanine, (S′PA). In contrast, the less conserved positions are enriched for serine encoded with AGY codons (denoted S″), glycine and asparagine, (GS″N). We tested this phenomenon in the HIV envelope glycoprotein (gp120), and the V-gene that encodes B-cell receptors/antibodies. These fast evolving proteins both have hypervariable positions, which are under diversifying selection, closely adjacent to highly conserved structural regions. In both instances, we identified an opposite abundance of two groups of serine substitutions, with enrichment of S′PA in the conserved positions, and GS″N in the hypervariable regions. Finally, we analyzed the substitutions across 60,000 individual human exomes to show that, when serine has a specific functional constraint of phosphorylation capability, S′ codons are 32-folds less prone than S″ to substitutions to Threonine or Tyrosine that could potentially retain the phosphorylation site capacity. Combined, our results, that cover evolutionary signals at different temporal scales, demonstrate that through its encoding by two codon sets, serine allows for the existence of alternating substitution patterns within positions of functional maintenance versus sites of rapid diversification.