WO2018185222A1 - Aminoacyl-arnt synthétases et arnt orthogonaux - Google Patents

Aminoacyl-arnt synthétases et arnt orthogonaux Download PDF

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WO2018185222A1
WO2018185222A1 PCT/EP2018/058731 EP2018058731W WO2018185222A1 WO 2018185222 A1 WO2018185222 A1 WO 2018185222A1 EP 2018058731 W EP2018058731 W EP 2018058731W WO 2018185222 A1 WO2018185222 A1 WO 2018185222A1
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trna
polynucleotide
seq
host cell
nucleotide sequence
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PCT/EP2018/058731
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Birgit Wiltschi
Tea PAVKOV KELLER
Patrik Fladischer
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Acib Gmbh - Austrian Centre Of Industrial Biotechnology
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/93—Ligases (6)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y601/00—Ligases forming carbon-oxygen bonds (6.1)
    • C12Y601/01—Ligases forming aminoacyl-tRNA and related compounds (6.1.1)
    • C12Y601/01026—Pyrrolysine-tRNAPyl ligase (6.1.1.26)

Definitions

  • the present invention relates to polynucleotides encoding polypeptides capable of catalyzing the aminoacylation of its cognate tRNA with aromatic or aliphatic amino acids to form an aminoacyl-tRNA, polypeptides encoded by such polynucleotides, vectors comprising such polynucleotides, host cells comprising such vectors, pairs of aminoacyl tRNA synthetases and cognate tRNAs, use of such compounds for catalyzing the aminoacylation of its cognate tRNA with aromatic or aliphatic amino acids to form an aminoacyl-tRNA, and methods for catalyzing the aminoacylation of its cognate tRNA with aromatic or aliphatic amino acids to form an aminoacyl-tRNA.
  • cAAs canonical amino acids
  • ncAAs offer a broad spectrum of side chain chemistries and many of them occur in nature (Walsh et al., Angewandte Chemie Int Ed (2013), 52 (28): 7098-7124).
  • aaRS aminoacyl-tRNA synthetase
  • SCS stop codon suppression
  • o-pair the pair consisting of the ncAA-specific aaRS and the tRNA C uA is orthogonal (o-pair), which means it does not interfere with the endogenous translation system of the host (Wang (2001 ), loc. cit.).
  • o-pairs developed were based on the archaeal TyrRS/tRNA C uA Tyr from Methanocaldococus janaschii (Mj) (Ryu et al., Nature Methods (2006), 3 (4): 263-265).
  • PylRS pyrrolysyl-tRNA synthetase
  • /WmPyIRS and ⁇ tePylRS are used routinely for the site-specific incorporation of a palette of ncAAs in E. coli.
  • the archaeal PylRS/tRNA C uA pairs are orthogonal not only in E. coli but also in other organisms, such as yeasts (Hancock et al., J Am Chem Soc (2010), 132 (42): 14819-14824) and mammalian cells (Mukai et al., Biochem Biophys Res Comm (2008), 371 (4): 818-822).
  • nucleotide sequence being at least 70%, 75%, 80%, 85%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the nucleotide sequence according to SEQ ID NO: 1 or 3 is codon-optimized to a selected host cell, and
  • tRNAs comprise an anticodon which are capable of identifying corresponding codons on the mRNA, usually either specific/base-by-base, or by wobble base pairing as known in the art. Also, as known in the art, usually tRNAs are specific for or prefer a particular amino acid with which the cognate tRNA may be charged by a corresponding aaRS. In one embodiment of the present invention, the amino acids which are charged onto the cognate tRNA are non-canonical.
  • Aromatic amino acids as used herein comprise canonical aromatic amino acids such as histidine, phenylalanine, tryptophan, and tyrosine, as well as non-canonical amino acids such as S-furyl-L-homocysteine, para-azido-L-phenylalanine, para-acetyl-L- phenylalanine, or para-propargyloxy-L-phenylalanine.
  • the tRNA corresponding to an aliphatic amino acid which are aminoacylated by the aaRS may be tRNA Pyl (tRNAcu A ), and the corresponding aliphatic amino acid may be Pyl or another lysine derivative.
  • a given enzyme may be considered to exhibit aminoacylation ability (i.e. ability to catalyze aminoacylation) if introduction (or incorporation, herein used synonymously in this context) of the corresponding amino acid into a polypeptide can be observed.
  • an amino acid into a polypeptide may be observed by any method known in the art, comprising inter alia mass spectrometry or Edman degradation as known in the art and also described and exemplified herein.
  • this aminoacylation process may preferably be catalyzed by polypeptides acting as aminoacyl tRNA synthetases as described and provided herein and encoded by the polynucleotides as described and provided herein. Said term may also be referred to herein as esterification of the cognate tRNA with an amino acid.
  • coli is capable of catalyzing aminoacylation of pyrrolysine (Pyl) with its cognate tRNA Pyl (tRNAcu A )-
  • the polypeptide encoded by the nucleotide sequence according to SEQ ID NO: 1 has an amino acid sequence as shown in SEQ ID NO: 2
  • the polypeptide encoded by the nucleotide sequence according to SEQ ID NO: 3 has an amino acid sequence as shown in SEQ ID NO: 4.
  • cognate tRNA as used in context with the present invention preferably means the tRNA which is charged or bonded with its corresponding amino acid to form an aminoacyl tRNA (also referred to herein as "aa tRNA”), a step which is preferably catalyzed by polypeptides encoded by polynucleotides as described and provided herein.
  • “Cognate” in this sense is understood by the person skilled in the art and may particularly mean a pair of aaRS and tRNA carrying the corresponding anticodon corresponding to the codon for said amino acid on the mRNA, either base-by-base or by wobble base pairing as known in the art.
  • a given aaRS recognizes a specific cognate tRNA and loads (charges, etc. as described herein) it with a corresponding amino acid.
  • a tRNA GA A (GAA being the anticodon of the tRNA) is charged with the corresponding amino acid phenylalanine (Phe) which is encoded on the mRNA by the codon UUC (or UUU).
  • the tRNA corresponding to an aliphatic amino acid which is aminoacylated by the aaRS encoded by a polynucleotide as described and provided herein may be a tRNA Pyl (tRNA C uA), and the corresponding aliphatic amino acid may be Pyl, and the aaRS may be a pyrrolysyl tRNA synthetase (PylRS).
  • Codon-optimization may also and additionally refer to exchange of different stop codons. For example, if a given host cell expresses certain suppressor molecules for certain stop codons, the stop codon (e.g., amber, ochre or opal) may be adapted accordingly. Such process as defined herein above is referred to herein as "codon-optimization".
  • codon usage tables which show the codon usage frequency for the respective host cell, i.e. which codons are used more often than others (and at which ratio).
  • codon usage tables are not available for all organisms (e.g., not for Candidates Methanomethylophilus alvus Mx1201 Ca). It is also one advantage of the present system that it is suitable to avoid interference with stop codons of the host cell while incorporating (preferably non-canonical) amino acids into polypeptides.
  • aminoacyl tRNA synthetase described and provided herein may be PylRS, which is able to aminoacylate a lysine derivative as described herein (e.g., pyrrolysine or boc-lysine) and its cognate tRNA Pyl (tRNA C uA)-
  • a lysine derivative as described herein e.g., pyrrolysine or boc-lysine
  • tRNA C uA tRNA Pyl
  • the anticodon CUA recognizes the amber stop codon (UAG) on the mRNA and, thus, acts as suppressor for the amber codon.
  • aaRS as described herein, wherein said nucleotide sequence is codon-optimized to a selected host cell which is not Candidatus Methanomethylophilus alvus Mx1201 Ca. That is, such aaRS according to the present invention are capable of catalyzing the aminoacylation of its cognate tRNA with an aliphatic or aromatic amino acid to form an aminoacyl-tRNA as described and exemplified herein.
  • aminoacyl tRNA synthetase may have an amino acid sequence which is at least 70%, 75%, 80%, 85%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 2, provided it is capable of catalyzing the aminoacylation of its cognate tRNA with an aliphatic or aromatic amino acid to form an aminoacyl-tRNA.
  • the aaRS of the present invention may also be a fragment, e.g.
  • the aaRS as described and provided herein may encompass 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid additions or substitutions compared to the amino acid sequence of SEQ ID NO: 2, preferably it may comprise 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10 conservative or highly conservative amino acid substitutions compared to the amino acid sequence of SEQ ID NO: 2.
  • silent mutations mean base substitutions within a nucleic acid sequence which do not change the amino acid sequence encoded by the nucleic acid sequence.
  • Constant mean substitutions as listed as “Exemplary Substitutions” in Table I below.
  • “Highly conservative” substitutions as used herein mean substitutions as shown under the heading "Preferred Substitutions" in Table I below.
  • position when used in accordance with the present invention means the position of an amino acid within an amino acid sequence depicted herein.
  • corresponding in this context also includes that a position is not only determined by the number of the preceding nucleotides/amino acids.
  • the level of identity between two or more sequences can be easily determined by methods known in the art, e.g., by BLAST analysis.
  • identity levels of nucleic acid sequences or amino acid sequences may refer to the entire length of the respective sequence and is preferably assessed pair-wise, wherein each gap is to be counted as one mismatch.
  • Deviations from the above-described nucleic acid sequences may have been produced, e.g., by deletion, substitution, addition, insertion and/or recombination.
  • the term “addition” refers to adding at least one nucleic acid residue/amino acid to the end of the given sequence, whereas "insertion” refers to inserting at least one nucleic acid residue/amino acid within a given sequence.
  • the term “deletion” refers to deleting or removal of at least one nucleic acid residue or amino acid residue in a given sequence.
  • substitution refers to the replacement of at least one nucleic acid residue/amino acid residue in a given sequence.
  • nucleic acid molecules may comprise inter alia DNA molecules, RNA molecules, oligonucleotide thiophosphates, substituted ribo-oligonucleotides or PNA molecules.
  • nucleic acid molecule may refer to DNA or RNA or hybrids thereof or any modification thereof that is known in the art (see, e.g., US 552571 1 , US 471 1955, US 5792608 or EP 302175 for examples of modifications).
  • the polynucleotide sequence may be single- or double- stranded, linear or circular, natural or synthetic, and without any size limitation.
  • the polynucleotide sequence may be genomic DNA, cDNA, mitochondrial DNA, mRNA, antisense RNA, ribozymal RNA or a DNA encoding such RNAs or chimeroplasts (Gamper, Nucleic Acids Research, 2000, 28, 4332 - 4339).
  • Said polynucleotide sequence may be in the form of a vector, plasmid or of viral DNA or RNA.
  • hybridization or “hybridizes” as used herein in context of nucleic acid molecules/DNA sequences may relate to hybridizations under stringent or non-stringent conditions. If not further specified, the conditions are preferably non-stringent. Said hybridization conditions may be established according to conventional protocols described, for example, in Sambrook, Russell “Molecular Cloning, A Laboratory Manual”, Cold Spring Harbor Laboratory, N. Y. (2001 ); Current Protocols in Molecular Biology, Update May 9, 2012, Print ISSN: 1934-3639, Online ISSN: 1934-3647; Ausubel, "Current Protocols in Molecular Biology", Green Publishing Associates and Wiley Interscience, N. Y.
  • Variations in the above conditions may be accomplished through the inclusion and/or substitution of alternate blocking reagents used to suppress background in hybridization experiments.
  • Typical blocking reagents include Denhardt's reagent, BLOTTO, heparin, denatured salmon sperm DNA, and commercially available proprietary formulations.
  • the inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility.
  • low stringent hybridization conditions for the detection of homologous or not exactly complementary sequences may, for example, be set at 6 x SSC, 1 % SDS at 65 °C.
  • the length of the probe and the composition of the nucleic acid to be determined constitute further parameters of the hybridization conditions.
  • Hybridizing nucleic acid molecules also comprise fragments of the above described molecules. Such fragments may represent nucleic acid molecules which code for a functional aaRS as described herein or a functional fragment thereof which can serve as a primer. Furthermore, nucleic acid molecules which hybridize with any of the aforementioned nucleic acid molecules also include complementary fragments, derivatives and variants of these molecules. Additionally, a hybridization complex refers to a complex between two nucleic acid sequences by virtue of the formation of hydrogen bonds between complementary G and C bases and between complementary A and T bases; these hydrogen bonds may be further stabilized by base stacking interactions. The two complementary nucleic acid sequences hydrogen bond in an antiparallel configuration.
  • hybridizing sequences preferably refers to sequences which display a sequence identity of at least 45%, more preferably at least 50%, more preferably at least 55%, more preferably at least 60%, more preferably at least 65%, more preferably at least 70%, more preferably at least 75%, more preferably at least 80%.
  • Any one or more of these silent mutations may serve as codon-optimization of the polynucleotide to any host cell which has a higher codon usage of CTG for Leu compared to CTC for Leu, and/or higher codon usage of CGT for Arg compared to AGG for Arg.
  • a non-exhaustive example for a host cell using more CTG than CTC for Leu and more CGT than AGG for Arg comprises E. coli.
  • said nucleotide sequence may comprise at least 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14, 15, 16, or 17 of any one of the following substitutions or pairs of substitutions a) to q), compared to the nucleotide sequence of SEQ ID NO: 1 :
  • polypeptide encoded by said polynucleotide is an aminoacyl tRNA synthetase (aaRS) capable of catalyzing the aminoacylation of its cognate tRNA with an aliphatic or aromatic amino acid to form an aminoacyl-tRNA as described and provided herein.
  • aaRS aminoacyl tRNA synthetase
  • the present invention also relates to a vector comprising a polynucleotide described and provided in accordance with the present invention. That is, the present invention also relates to a vector comprising a polynucleotide as described and provided herein encoding an aminoacyl tRNA synthetase (aaRS) capable of catalyzing the aminoacylation of its cognate tRNA with an aliphatic or aromatic amino acid to form an aminoacyl-tRNA as described and also provided herein, said polynucleotide comprising a nucleotide sequence which is at least 70%, 75%, 80%, 85%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the nucleotide sequence according to SEQ ID NO: 1 or 3, wherein said nucleotide sequence is codon-optimized to a selected host cell which is not Candidates Methanomethylophilus alvus Mx1201 Ca.
  • aaRS aminoacyl t
  • the amino acid which is charged to the corresponding tRNA by the aaRS e.g., PylRS, for example the PylRS from Mma according to SEQ ID NO: 2 as described herein is a pyrrolysine or boc-lysine, or AllocK, or AzideK, or another pyrrolysine derivative as described herein
  • the tRNA may be tRNA Pyl (e.g., MmatRNA Pyl according to SEQ ID NO: 5 to have the orthogonal Mma-pair (MmaOP), or /WmtRNA Pyl derived from Mm to have a hybrid pair).
  • the nucleic acid construct is preferably inserted into that vector in a manner the resulting vector comprises only one promoter suitable to be employed in context of this invention.
  • the promoter can be excised either from the nucleic acid construct or from the expression vector prior to ligation.
  • the vector is able to integrate into the host cell genome.
  • the vector may be any vector suitable for the respective host cell, preferably an expression vector.
  • the present invention also relates to an aminoacyl tRNA synthetase (aaRS) as described and provided herein, and to a tRNA as described and provided herein.
  • the aminoacyl tRNA synthetase may be a pyrrolysine tRNA synthetase (PylRS), for example the PylRS derived from Candidatus Methanomethylophilus alvus Mx1201 Ca.
  • Lysogen broth (LB) medium (Roth) was used for all stop codon suppression experiments.
  • E. coli BL21 cells harboring a pSCS+ plasmid carrying an o-pair consisting of a PylRS and a tRNA C uA were cultivated in 250 mL flasks each containing 50 mL LB medium with 50 ⁇ g/mL kanamycin (Roth).
  • the initial cell density D 600 was 0.1. Cultures were incubated at 37 °C on an orbital shaker at 160 rpm. At D 600 of 0.8-1.0, the expression of the PylRS was induced by adding 0.2% (w/v) of arabinose (Roth).
  • Cells were diluted 1 :5 (v/v) with 1 x PBS buffer (137 mM NaCI, 2.7 mM KCI, 10 mM Na 2 HP0 4 , 2 mM KH 2 P0 4 ) and 8 times 100 ⁇ _ were measured in a NUNC flat bottom 96-well black plate (Thermo Fisher Scinetific) after 5 seconds of vigorous shaking.
  • SDS-PAGE using 4-12% Bis-Tris SDS-gels was performed following the instructions of the manufacturer.

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Abstract

La présente invention concerne des polynucléotides capables de catalyser l'aminoacylation de leur ARNt apparenté avec des acides aminés aromatiques ou aliphatiques pour former un aminoacyl-ARNt, des polypeptides codés par de tels polynucléotides, des vecteurs comprenant de tels polynucléotides, des cellules hôtes comprenant de tels vecteurs, des paires d'aminoacyl-ARNt synthétases et d'ARNt apparentés, l'utilisation de tels composés pour catalyser l'aminoacylation d'acides aminés aromatiques ou aliphatiques avec l'ARNt correspondant pour former un aminoacyl-ARNt, et des procédés pour catalyser l'aminoacylation d'ARNt apparenté avec des acides aminés aromatiques ou aliphatiques pour former un aminoacyl-ARNt.
PCT/EP2018/058731 2017-04-05 2018-04-05 Aminoacyl-arnt synthétases et arnt orthogonaux WO2018185222A1 (fr)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20210324363A1 (en) * 2018-08-31 2021-10-21 Riken Pyrrolysyl-trna synthetase
WO2022003142A1 (fr) 2020-07-03 2022-01-06 Engenes Biotech Gmbh Variants de pyrrolysyl-arnt synthétase et leurs utilisations
WO2023031445A3 (fr) * 2021-09-06 2023-04-13 Veraxa Biotech Gmbh Nouveaux variants d'aminoacyl-arnt synthétase pour l'expansion de code génétique dans des eucaryotes

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4711955A (en) 1981-04-17 1987-12-08 Yale University Modified nucleotides and methods of preparing and using same
EP0302175A2 (fr) 1982-06-23 1989-02-08 Enzo Biochem, Inc. Nucléotides et polynucléotides modifiés marqués et méthodes pour leur préparation, utilisation et détection
US5525711A (en) 1994-05-18 1996-06-11 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Pteridine nucleotide analogs as fluorescent DNA probes
US5792608A (en) 1991-12-12 1998-08-11 Gilead Sciences, Inc. Nuclease stable and binding competent oligomers and methods for their use
WO2014082773A1 (fr) 2012-11-30 2014-06-05 Universite D'auvergne Clermont I Utilisation de microorganismes pour diminuer le taux de triméthylamine dans une cavité du corps humain, notamment pour le traitement de la triméthylaminurie ou d'une vaginose bactérienne et la prévention des maladies cardiovasculaires

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4711955A (en) 1981-04-17 1987-12-08 Yale University Modified nucleotides and methods of preparing and using same
EP0302175A2 (fr) 1982-06-23 1989-02-08 Enzo Biochem, Inc. Nucléotides et polynucléotides modifiés marqués et méthodes pour leur préparation, utilisation et détection
US5792608A (en) 1991-12-12 1998-08-11 Gilead Sciences, Inc. Nuclease stable and binding competent oligomers and methods for their use
US5525711A (en) 1994-05-18 1996-06-11 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Pteridine nucleotide analogs as fluorescent DNA probes
WO2014082773A1 (fr) 2012-11-30 2014-06-05 Universite D'auvergne Clermont I Utilisation de microorganismes pour diminuer le taux de triméthylamine dans une cavité du corps humain, notamment pour le traitement de la triméthylaminurie ou d'une vaginose bactérienne et la prévention des maladies cardiovasculaires
US20160074440A1 (en) * 2012-11-30 2016-03-17 Universite D'auvergne Clermont I Use of microorganisms for reducing the level of trimethylamine in a human body cavity, in particular for the treatment of trimethylaminuria or of bacterial vaginosis and the prevention of cardiovascular diseases

Non-Patent Citations (45)

* Cited by examiner, † Cited by third party
Title
"Current Protocols in Molecular Biology", 9 May 2012
"Nucleic acid hybridization, a practical approach", 1985, IRL PRESS
AUSUBEL: "Current Protocols in Molecular Biology", 1989, GREEN PUBLISHING ASSOCIATES AND WILEY INTERSCIENCE
DATABASE EMBL [online] 10 April 2013 (2013-04-10), "Candidatus Methanomethylophilus alvus Mx1201 Pyrrolysyl-tRNA synthetase", XP002781482, retrieved from EBI accession no. EMBL:AGI85861 Database accession no. AGI85861 *
DATABASE ENA [online] 15 May 2014 (2014-05-15), "Thermoplasmatales archaeon BRNA1, complete genomeCP002916; SV 1; circular; genomic DNA; Thermoplasmatales archaeon BRNA1, complete genome", XP002772180, Database accession no. CP002916 *
DATABASE GSN [online] 31 July 2014 (2014-07-31), "Methanomethylophilus alvus genomic DNA, SEQ ID 13", XP002772178, Database accession no. BBH10200 *
DATABASE UniProt [online] 13 April 2016 (2016-04-13), "SubName: Full=Pyrrolysine--tRNA ligase PylS {ECO:0000313|EMBL:AMH95040.1};", XP002772183, retrieved from EBI accession no. UNIPROT:A0A110A2Y8 Database accession no. A0A110A2Y8 *
DATABASE UniProt [online] 15 March 2017 (2017-03-15), "SubName: Full=Pyrrolysyl-tRNA synthetase {ECO:0000313|EMBL:AGI85861.1}; EC=6.1.1.26 {ECO:0000313|EMBL:AGI85861.1};", XP002772179, retrieved from EBI accession no. UNIPROT:M9SC49 Database accession no. M9SC49 *
DATABASE UniProt [online] 15 March 2017 (2017-03-15), "SubName: Full=Seryl-tRNA synthetase {ECO:0000313|EMBL:AGI48081.1};", XP002772182, retrieved from EBI accession no. UNIPROT:M4YUJ0 Database accession no. M4YUJ0 *
DATABASE UniProt [online] 2 November 2016 (2016-11-02), "SubName: Full=Uncharacterized protein {ECO:0000313|EMBL:KUE74102.1};", XP002772181, retrieved from EBI accession no. UNIPROT:A0A0W7TK26 Database accession no. A0A0W7TK26 *
DAVID G. LONGSTAFF ET AL: "A natural genetic code expansion cassette enables transmissible biosynthesis and genetic encoding of pyrrolysine", PROCEEDINGS NATIONAL ACADEMY OF SCIENCES PNAS, vol. 104, no. 3, 4 January 2007 (2007-01-04), US, pages 1021 - 1026, XP055391157, ISSN: 0027-8424, DOI: 10.1073/pnas.0610294104 *
DUMAS ET AL., CHEMICAL SCIENCE, vol. 6, no. 1, 2015, pages 50 - 59
FRANCKLYN ET AL.: "Methods for kinetic and thermodynamic analysis of aminoacyl-tRNA synthetases", METHODS, vol. 44, no. 2, 2008, pages 100 - 118, XP022437407, DOI: doi:10.1016/j.ymeth.2007.09.007
G. BORREL ET AL: "Genome Sequence of "Candidatus Methanomethylophilus alvus" Mx1201, a Methanogenic Archaeon from the Human Gut Belonging to a Seventh Order of Methanogens", JOURNAL OF BACTERIOLOGY, vol. 194, no. 24, 15 December 2012 (2012-12-15), US, pages 6944 - 6945, XP055391266, ISSN: 0021-9193, DOI: 10.1128/JB.01867-12 *
GAMPER, NUCLEIC ACIDS RESEARCH, vol. 28, 2000, pages 4332 - 4339
GASTON ET AL., CURR OP MICROBIOL, vol. 14, no. 3, 2011, pages 342 - 349
GUILLAUME BORREL ET AL: "Comparative genomics highlights the unique biology of Methanomassiliicoccales, a Thermoplasmatales-related seventh order of methanogenic archaea that encodes pyrrolysine", BMC GENOMICS, vol. 15, no. 1, 679, 13 August 2014 (2014-08-13), BIOMED CENTRAL LTD, LONDON, UK, pages 1 - 23, XP021195522, ISSN: 1471-2164, DOI: 10.1186/1471-2164-15-679 *
GUILLAUME BORREL ET AL: "Supplementary data: Unique Characteristics of the Pyrrolysine System in the 7th Order of Methanogens: Implications for the Evolution of a Genetic Code Expansion Cassette", ARCHAEA, 27 January 2014 (2014-01-27), pages 1 - 4, XP055391263, Retrieved from the Internet <URL:http://downloads.hindawi.com/journals/archaea/2014/374146.f1.pdf> [retrieved on 20170717], DOI: 10.1155/2014/374146 *
GUILLAUME BORREL ET AL: "Unique Characteristics of the Pyrrolysine System in the 7th Order of Methanogens: Implications for the Evolution of a Genetic Code Expansion Cassette", ARCHAEA, vol. 2014, 27 January 2014 (2014-01-27), CA, pages 1 - 11, XP055391251, ISSN: 1472-3646, DOI: 10.1155/2014/374146 *
HANCOCK ET AL., J AM CHEM SOC, vol. 132, no. 42, 2010, pages 14819 - 14824
HERRING ET AL., NUCLEIC ACID RES, vol. 35, no. 4, 2007, pages 1270 - 1278
HOHSAKA ET AL., J BIOL CHEM, vol. 6, no. 6, 2010, pages 809 - 815
KATAYAMA ET AL., BIOSCIENCE, BIOTECHNOL, BIOCHEM, vol. 76, no. 1, 2012, pages 205 - 208
KAVRAN ET AL., PNAS USA, vol. 104, no. 27, 2007, pages 11268 - 11273
KRZYCKI ET AL., CURR OP MICROBIOL, vol. 8, no. 6, 2005, pages 706 - 712
LIU ET AL., PNAS, vol. 94, 1997, pages 10092 - 10097
MUKAI ET AL., BIOCHEM BIOPHYS RES COMM, vol. 371, no. 4, 2008, pages 818 - 822
NAMY ET AL: "Adding pyrrolysine to the Escherichia coli genetic code", FEBS LETTERS, vol. 581, no. 27, 23 October 2007 (2007-10-23), ELSEVIER, AMSTERDAM, NL, pages 5282 - 5288, XP022324692, ISSN: 0014-5793, DOI: 10.1016/J.FEBSLET.2007.10.022 *
NOZAWA ET AL., NATURE, vol. 457, no. 7233, 2009, pages 1163 - 1167
POLYCARPO ET AL., FEBS LETTERS, vol. 580, no. 28-29, 2006, pages 6695 - 6670
RYU ET AL., NATURE METHODS, vol. 3, no. 4, 2006, pages 263 - 265
SAMBROOK; RUSSELL: "Molecular Cloning, A Laboratory Manual", 2001, COLD SPRING HARBOR LABORATORY
SCHRADER ET AL., PNAS, vol. 108, no. 13, 2011, pages 5215 - 5220
SHEVCHENKO, ANALYTICAL CHEMISTRY, vol. 68, no. 5, 1996, pages 850 - 858
SRINIVASAN ET AL., SCIENCE, vol. 296, no. 5572, 2002, pages 1459 - 1462
UTTAMAPINAT ET AL., ANGEW CHEM INT ED, vol. 51, no. 24, 2012, pages 5852 - 5856
WALSH ET AL., ANGEWANDTE CHEMIE INT ED, vol. 52, no. 28, 2013, pages 7098 - 7124
WAN ET AL., BIOCHIMICA ET BIOPHYSICA ACTA (BBA) - PROTEINS AND PROTEOMICS, vol. 1844, no. 6, 2014, pages 1059 - 1070
WANG ET AL., MOL BIOSYSTEMS, vol. 7, no. 3, 2011, pages 714 - 717
WANG ET AL., SCIENCE, vol. 292, no. 5516, 2001, pages 498 - 500
WANG; SCHULTZ, CHEM BIOL, vol. 8, 2001, pages 883 - 890
YANAGISAWA ET AL., CHEM & BIOL, vol. 15, no. 11, 2008, pages 1187 - 1197
YANE-SHIH WANG ET AL: "Supplementary Material: The de novo engineering of pyrrolysyl-tRNA synthetase for genetic incorporation of l-phenylalanine and its derivatives", MOLECULAR BIOSYSTEMS, vol. 7, no. 3, 14 January 2011 (2011-01-14), GB, pages S1 - S21+11PP, XP055391381, ISSN: 1742-206X, DOI: 10.1039/c0mb00217h *
YANE-SHIH WANG ET AL: "The de novo engineering of pyrrolysyl-tRNA synthetase for genetic incorporation of l-phenylalanine and its derivatives", MOLECULAR BIOSYSTEMS, vol. 7, no. 3, 14 January 2011 (2011-01-14), GB, pages 714 - 717, XP055391379, ISSN: 1742-206X, DOI: 10.1039/c0mb00217h *
YOUNG ET AL., J BIOL CHEM, vol. 285, no. 15, 2010, pages 11039 - 11044

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20210324363A1 (en) * 2018-08-31 2021-10-21 Riken Pyrrolysyl-trna synthetase
WO2022003142A1 (fr) 2020-07-03 2022-01-06 Engenes Biotech Gmbh Variants de pyrrolysyl-arnt synthétase et leurs utilisations
WO2023031445A3 (fr) * 2021-09-06 2023-04-13 Veraxa Biotech Gmbh Nouveaux variants d'aminoacyl-arnt synthétase pour l'expansion de code génétique dans des eucaryotes

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