An unnatural base pair for incorporating amino acid analogs into proteins

  title={An unnatural base pair for incorporating amino acid analogs into proteins},
  author={Ichiro Hirao and Takashi Ohtsuki and Tsuyoshi Fujiwara and Tsuneo Mitsui and Tomoko Yokogawa and Taeko Okuni and Hiroshi Nakayama and Koji Takio and Takashi Yabuki and Takanori Kigawa and Koichiro Kodama and Takashi Yokogawa and Kazuya Nishikawa and Shigeyuki Yokoyama},
  journal={Nature Biotechnology},
An unnatural base pair of 2-amino-6-(2-thienyl)purine (denoted by s) and pyridin-2-one (denoted by y) was developed to expand the genetic code. The ribonucleoside triphosphate of y was site-specifically incorporated into RNA, opposite s in a template, by T7 RNA polymerase. This transcription was coupled with translation in an Escherichia coli cell-free system. The yAG codon in the transcribed ras mRNA was recognized by the CUs anticodon of a yeast tyrosine transfer RNA (tRNA) variant, which had… 
Site-Specific Incorporation of Functional Components into RNA by an Unnatural Base Pair Transcription System
It is found that the efficient incorporation into a position close to the 3′-terminus of a transcript depended on the natural base contexts neighboring the unnatural base, and that pyrimidine-Ds-pyrimidine sequences in templates were generally favorable, relative to purine-Ds -purine sequences.
Site-specific incorporation of an unnatural amino acid into proteins in mammalian cells.
It is found that amber suppression can occur with the heterologous pair of E.coli TyrRS and Bacillus stearothermophilus suppressor tRNA(Tyr), which naturally contains the promoter sequence.
Phosphoserine aminoacylation of tRNA bearing an unnatural base anticodon.
A linear DNA template-based framework for site-specific unnatural amino acid incorporation
An unnatural hydrophobic base pair system: site-specific incorporation of nucleotide analogs into DNA and RNA
An unnatural base Pair system in which DNA containing an unnatural base pair can be amplified and function as a template for the site-specific incorporation of base analog substrates into RNA via transcription is presented.
Site-specific biotinylation of RNA molecules by transcription using unnatural base pairs
Direct site-specific biotinylation of RNA molecules was achieved by specific transcription mediated by unnatural base pairs applied to the immobilization of an RNA aptamer on sensor chips and the aptamer accurately recognized its target protein.
PCR Amplification and Transcription for Site-Specific Labeling of Large RNA Molecules by a Two-Unnatural-Base-Pair System
A PCR and transcription system using two hydrophobic unnatural base pairs, combining the Ds-Px and D's-Pa pairs with modified Pa substrates, provides a powerful tool for the site-specific labeling and modification of desired positions in large RNA molecules.
Ribosomal synthesis of unnatural peptides.
It is shown that the chemical space sampled byinatorial libraries of non-biological polymers and drug-like peptides can be expanded using mutant aminoacyl-tRNA synthetases for the incorporation of additional unnatural amino acids or by the specific posttranslational chemical derivitization of reactive groups with small molecules.


Unnatural base pairs for specific transcription
The bulky dimethylamino group of x in the templates effectively eliminates noncognate pairing with the natural bases and enables RNA biosynthesis for the specific incorporation of unnatural nucleotides at the desired positions.
A general method for site-specific incorporation of unnatural amino acids into proteins.
The ability to selectively replace amino acids in a protein with a wide variety of structural and electronic variants should provide a more detailed understanding of protein structure and function.
Effects of release factor 1 on in vitro protein translation and the elaboration of proteins containing unnatural amino acids.
An in vitro protein synthesizing system was modified to facilitate the improved, site-specific incorporation of unnatural amino acids into proteins via readthrough of mRNA nonsense (UAG) codons by chemically misacylated suppressor tRNAs, resulting in a substantial increase in mutant protein yields.
An Engineered Tetrahymena tRNAGln for in Vivo Incorporation of Unnatural Amino Acids into Proteins by Nonsense Suppression*
A new tRNA, THG73, has been designed and evaluated as a vehicle for incorporating unnatural amino acids site-specifically into proteins expressed in vivo using the stop codon suppression technique and is as much as 100-fold less likely to be acylated by endogenous synthetases of the Xenopus oocyte.
Efficient Incorporation of Nonnatural Amino Acids with Large Aromatic Groups into Streptavidin in In Vitro Protein Synthesizing Systems
Efficiencies of the incorporation of various nonnatural amino acids carrying aromatic side groups into streptavidin were examined and indicated that the efficiencies were higher for aromatic groups with straight configurations than those with widely expanded or bend configurations.
Efficient incorporation of unsaturated methionine analogues into proteins in vivo
Results of the in vitro assays corroborate the in vivo incorporation results, suggesting that success or failure of analogue incorporation in vivo is controlled by MetRS.
Changing the amino acid specificity of yeast tyrosyl-tRNA synthetase by genetic engineering.
This mutant TyrRS should serve as a new tool for site-specific incorporation of non-canonical amino acids, such as those in 3-substituted tyrosine analogues, into proteins in an appropriate translation system in vivo or in vitro.
Expanding the Genetic Code of Escherichia coli
A unique transfer RNA/aminoacyl-tRNA synthetase pair has been generated that expands the number of genetically encoded amino acids in Escherichia coli and should provide a general method for increasing the genetic repertoire of living cells to include a variety of amino acids with novel structural, chemical, and physical properties not found in the common 20 amino acids.