Probing the Limits of Genetic Recoding in Essential Genes

@article{Lajoie2013ProbingTL,
  title={Probing the Limits of Genetic Recoding in Essential Genes},
  author={Marc J. Lajoie and Sriram Kosuri and Joshua A. Mosberg and Christopher J. Gregg and D. Zhang and George M. Church},
  journal={Science},
  year={2013},
  volume={342},
  pages={361 - 363}
}
Changing the Code Easily and efficiently expanding the genetic code could provide tools to genome engineers with broad applications in medicine, energy, agriculture, and environmental safety. Lajoie et al. (p. 357) replaced all known UAG stop codons with synonymous UAA stop codons in Escherichia coli MG1655, as well as release factor 1 (RF1; terminates translation at UAG), thereby eliminating natural UAG translation function without impairing fitness. This made it possible to reassign UAG as a… 
Genomically Recoded Organisms Expand Biological Functions
TLDR
The construction and characterization of a genomically recoded organism (GRO) is described, which exhibited improved properties for incorporation of nonstandard amino acids that expand the chemical diversity of proteins in vivo and exhibited increased resistance to T7 bacteriophage, demonstrating that new genetic codes could enable increased viral resistance.
Organisms with alternative genetic codes resolve unassigned codons via mistranslation and ribosomal rescue
TLDR
Recoded organisms resolve translation at unassigned UAG codons via near-cognate suppression, dramatic frameshifting from at least −3 to +19 nucleotides, and rescue by ssrA-encoded tmRNA, ArfA, and ArfB, demonstrating that genomic recoding is a promising path for impairing horizontal gene transfer and conferring genetic isolation in diverse organisms.
Revealing the amino acid composition of proteins within an expanded genetic code
TLDR
A new proteomic workflow was created that enabled the detection of UAG readthrough in native proteins in E. coli strains in which UAG was reassigned to encode phosphoserine and quantitation of NSAA and natural AA incorporation at UAG in a recombinant reporter protein.
Design, synthesis, and testing toward a 57-codon genome
TLDR
Computational design, synthesis, and progress toward assembly of a 3.97-megabase, 57-codon Escherichia coli genome in which all 62,214 instances of seven codons were replaced with synonymous alternatives across all protein-coding genes are reported.
Emergent rules for codon choice elucidated by editing rare arginine codons in Escherichia coli
Significance This work presents the genome-wide replacement of all rare AGR (AGA and AGG) arginine codons in the essential genes of Escherichia coli with synonymous CGN alternatives. Synonymous codon
Efficient Reassignment of a Frequent Serine Codon in Wild-Type Escherichia coli.
TLDR
The results indicate that the 3-iodo-l-phenylalanyl-tRNA synthetase (IFRS)/tRNA(Pyl) pair can efficiently outcompete the cellular machinery to reassign select sense codons in wild-type E. coli.
Reassignment of a rare sense codon to a non-canonical amino acid in Escherichia coli
TLDR
The in vivo reassignment of the AGG sense codon from arginine to l-homoarginine showed the feasibility of breaking the degeneracy of sense codons to enhance the amino-acid diversity in the genetic code.
Stop Codon Reassignment in the Wild
TLDR
This work carried out a systematic analysis of stop codon reassignments from the canonical TAG amber, TGA opal, and TAA ochre codons in assembled metagenomes from environmental and host-associated samples, single-cell genomes of uncultivated bacteria and archaea, and a collection of phage sequences.
Future of the Genetic Code
TLDR
Evidence suggests that the problem of anticodons as identity elements can be diminished or resolved through removal from the tRNA of all identity elements besides the anticodon, and theproblem of misreading of NNY codons by UNN anticodon can be resolved by the retirement of both the UNN and its complementary NNA codon from the proteome.
Next-generation genetic code expansion.
...
...

References

SHOWING 1-10 OF 59 REFERENCES
Genomically Recoded Organisms Expand Biological Functions
TLDR
The construction and characterization of a genomically recoded organism (GRO) is described, which exhibited improved properties for incorporation of nonstandard amino acids that expand the chemical diversity of proteins in vivo and exhibited increased resistance to T7 bacteriophage, demonstrating that new genetic codes could enable increased viral resistance.
Natural expansion of the genetic code.
TLDR
The existence of phosphoseryl-tRNA (in the form of tRNACys and tRNASec) may presage the discovery of other cotranslationally inserted modified amino acids.
Improving Lambda Red Genome Engineering in Escherichia coli via Rational Removal of Endogenous Nucleases
TLDR
Removing a set of five exonucleases substantially improves the performance of co-selection multiplex automatable genome engineering (CoS-MAGE) and investigates and clarify the effects of oligonucleotide phosphorothioation on recombination frequency.
Precise Manipulation of Chromosomes in Vivo Enables Genome-Wide Codon Replacement
TLDR
H hierarchical conjugative assembly genome engineering (CAGE) was developed to merge these sets of codon modifications into genomes with 80 precise changes, which demonstrate that these synonymous codon substitutions can be combined into higher-order strains without synthetic lethal effects.
Synonymous codon usage in Drosophila melanogaster: natural selection and translational accuracy.
TLDR
Evidence is presented that natural selection biases synonymous codon usage to enhance the accuracy of protein synthesis in Drosophila melanogaster to avoid translational misincorporation and to support functional constraint at the protein level.
Determinants of translation efficiency and accuracy
TLDR
New means to model the process of translation in a richer framework that will incorporate information about gene sequences, the tRNA pool of the organism and the thermodynamic stability of the mRNA transcripts are suggested.
The anti-Shine-Dalgarno sequence drives translational pausing and codon choice in bacteria
TLDR
It is shown that internal SD-like sequences are a major determinant of translation rates and a global driving force for the coding of bacterial genomes, and that codons decoded by rare transfer RNAs do not lead to slow translation under nutrient-rich conditions.
One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products.
  • K. Datsenko, B. Wanner
  • Biology, Engineering
    Proceedings of the National Academy of Sciences of the United States of America
  • 2000
TLDR
A simple and highly efficient method to disrupt chromosomal genes in Escherichia coli in which PCR primers provide the homology to the targeted gene(s), which should be widely useful, especially in genome analysis of E. coli and other bacteria.
Creation of a Bacterial Cell Controlled by a Chemically Synthesized Genome
TLDR
The design, synthesis, and assembly of the 1.08–mega–base pair Mycoplasma mycoides JCVI-syn1.0 genome starting from digitized genome sequence information and its transplantation into a M. capricolum recipient cell to create new cells that are controlled only by the synthetic chromosome are reported.
...
...