Riken Corporation

Japan Science and Technology Agency

Kyowa Kirin Co., Ltd.

Japan Science and Technology Corporation

Riken Japan Science and Technology Agency of 1-8

Kensaku Sakamoto

Shigeyuki Yokoyama

Takahito Mukai

Takatsugu Kobayashi

Tatsuo Yanagisawa

Akiko Matsumoto

Yasuhisa Shiraishi

Ichiro Hirao

Mikako Shirouzu

Akifumi Kato

Kaname Kimura

Masakazu Homma

Yoshimi Yamaguchi

Nobumasa Hino

Fumie Iraha

Kazumasa Ohtake

Daisuke Kiga

Ayako Sakamoto

Kenji Oki

The present invention relates to a monoclonal antibody or antibody fragment thereof as a human IgG antibody including at least one lysine derivative in a constant region of the human IgG antibody; a modified antibody or antibody fragment thereof, wherein the lysine derivative is modified; a nucleic acid including a nucleotide sequence encoding the antibody or antibody fragment thereof; a vector including the nucleic acid; a transformed cell obtained by introducing the vector into a host cell; a method for producing the antibody or antibody fragment thereof; and a composition containing the antibody or antibody fragment thereof.

Antibody with non-natural amino acid introduced therein

The present invention provides a novel method of producing a recombinant bacterium for production of a non-natural protein, including: (1) expressing tRNA in a bacterium, which tRNA recognizes UAG codon; (2) expressing an aminoacyl-tRNA synthetase in the bacterium, which aminoacyl-tRNA synthetase acylates the tRNA with a non-natural amino acid or an &#x3b1;-hydroxy acid; (3) (i) introducing a DNA construct into the bacterium, which DNA construct is for expressing, in the absence of a release factor for terminating translation at UAG codon, a function of at least one gene selected from the group consisting of genes each of which loses its function when a gene that codes for the release factor is defective and/or introducing an alteration into said at least one gene in a chromosome of the bacterium, which alteration is for expressing the function of said at least one gene in the absence of the release factor; and (4) causing the gene that codes for the release factor in the bacterium to be defective.

Method for constructing recombinant bacterium for producing non-native protein, and utilization of same

Method for incorporating a lysine derivative (particularly an N-benzyloxycarbonyl-lysine (Z-Lys) derivative) having useful functional group such as heavy atom, selenium, reactive functional group, fluorescent group or crosslinker, which is suitable as a non-natural amino acid, into a desired protein in a site-specific manner. A mutant pyrrolysyl-tRNA synthetase has substitution of at least one amino acid residue selected from tyrosine residue at position 306, leucine residue at position 309 and cysteine residue at position 348 each constituting a pyrrolysine-binding site in the amino acid sequence for pyrrolysyl-tRNA synthetase of SEQ ID NO:2. The substitution of the amino acid residue is: of tyrosine residue at position 306 by glycine or alanine residue, of leucine residue at position 309 by glycine or alanine residue, and/or of a cysteine residue at position 348 by valine, serine or alanine residue.

Mutant pyrrolysyl-tRNA synthetase, and method for production of protein having non-natural amino acid integrated therein by using the same

There are provided a DNA construct comprising a suppressor tRNA gene of a non-eukaryote containing no internal promoter functioning in a eukaryotic cell, and a eukaryotic or bacteriophage promoter linked at the 5' end of the tRNA gene, a method for synthesizing a suppressor tRNA by using the DNA construct, and a process for producing protein incorporating a non-natural amino acid by using the same.

Method of synthesizing a suppressor tRNA, DNA construct and use thereof for producing a non-natural amino acid-incorporated protein

A non-natural protein having at least one ester bond in its polypeptide main chain is synthesized by using an in vivo translation system in a ribosome. The following components (a) to (c) are expressed in a cell or an cell extraction solution in the presence of an &#x3b1;-hydroxy acid: (a) an aminoacyl-tRNA synthetase which can activate the &#x3b1;-hydroxy acid; (b) suppressor tRNA which can bind to the &#x3b1;-hydroxy acid in the presence of the aminoacyl-tRNA synthetase; and (c) a gene encoding a desired protein having a nonsense mutation or a frame-shift mutation at a desired site.

Process for production of non-natural protein having ester bond therein

Producing proteins incorporating non-natural amino acids can involve introducing genes into and knocking inherent genes out of eukaryote-type cells. Genes to be introduced include genes encoding eukaryote-type aminoacyl tRNA synthetase mutants having enhanced specificity to non-natural amino acids, compared with specificity to similar natural amino acids, and tRNA genes for non-natural amino acids capable of binding to the non-natural amino acids in the presence of the eukaryote-type aminoacyl tRNA synthetase mutants. Inherent genes to be knocked out include genes encoding aminoacyl tRNA synthetase having specificity to natural amino acids and tRNA genes capable of binding to the natural amino acids in the presence of the inherent aminoacyl tRNA synthetase.

Method for producing proteins comprising non-natural amino acids incorporated therein

A polypeptide according to the present invention includes: an altered polypeptide obtained by altering an ArgRS, a CysRS, a MetRS, a GlnRS, a GluRS, a LysRS, a TyrRS, or a TrpRS so that an unnatural amino acid is recognized; and an editing polypeptide derived from a PheRS, a LeuRS, an IleRS, a ValRS, an AlaRS, a ProRS, or a ThrRS, the editing polypeptide having been either inserted between a Rossman-fold N domain and a Rossman-fold C domain that exist in the altered polypeptide, or bound to an N terminal of the altered polypeptide. Thus provided are a new aaRS that exhibits high substrate specificity to an unnatural amino acid and a technique that involves the use of such an aaRS.

Polypeptide having activity of aminoacyl-tRNA synthetase and use thereof

This invention provides a method for producing proteins into which non-natural amino acids have been incorporated at desired positions involving the use of either prokaryote-derived or eukaryote-type aaRS* and prokaryotic cells or eukaryote-type as host cells, the method comprising steps of: (a) introducing genes encoding prokaryote-derived aminoacyl tRNA synthetase mutants having enhanced specificity to non-natural amino acids similar to given natural amino acids (compared with specificity to the natural amino acids) and tRNA genes for non-natural amino acids capable of binding to the non-natural amino acids in the presence of the prokaryote-derived aminoacyl tRNA synthetase mutants into prokaryotic cells that express genes encoding eukaryote-type aminoacyl tRNA synthetase having specificity to the given natural amino acids and tRNA genes capable of binding to the natural amino acids in the presence of eukaryote-type aminoacyl tRNA synthetase; and (b) knocking out genes encoding aminoacyl tRNA synthetase having specificity to the natural amino acids, which are inherent in the prokaryotic cells, and inherent tRNA genes capable of binding to the natural amino acids in the presence of the inherent aminoacyl tRNA synthetase.

Method for production of protein having non-natural type amino acid integrated therein

There are provided a DNA construct comprising non-eukaryote-derived suppressor tRNA gene containing no internal promoter functioning in a eukaryotic cell, and a eukaryote-derived or bacteriophage-derived promoter linked at the 5' end of the tRNA gene, a method for synthesizing a suppressor tRNA by using the DNA construct, and a process for producing a non-natural amino acid-incorporated protein by using the same.

Method of synthesizing a suppressor TRNA, DNA construct and use thereof for producing a protein including a non-natural amino acid

The present invention discloses an expression method for non-naturally-occurring amino acid-containing protein comprising: expressing in animal cells: (A) a mutant tyrosyl-tRNA synthetase that is a mutation of tyrosyl-tRNA synthetase originating inwith an enhanced specificity for a non-naturally-occurring tyrosine derivative as compared with the specificity for tyrosine; (B) suppressor tRNA originating inspecies,species orspecies of eubacteria and capable of binding with the tyrosine derivative in the presence of the mutant tyrosyl tRNA synthetase; and, (C) a desired protein gene that has undergone a nonsense mutation at a desired site; wherein, the tyrosine derivative is incorporated at the location of this nonsense mutation.

DNA sequences for the expression of alloproteins

A method of expressing a protein having an unnatural amino acid integrated thereinto comprising expressing (A) a mutant tyrosyl tRNA synthase that is derived from-origin tyrosyl tRNA synthase and has an elevated specificity for an unnatural tyrosine derivative compared with the specificity for tyrosine, (B) a suppressor tRNA originating in an eubacterium belonging to the genusorwhich is capable of binding to the above tyrosine derivative in the presence of the above mutant TyrRS, and (C) a desired protein gene having a nonsense mutation at a desired site in animal cells to thereby incorporate the above tyrosine derivative into the nonsense mutation site in the above protein.

Method of expressing proteins comprising non-naturally-occurring amino acids

Tyrosyl-tRNA synthetases having a modified amino acid sequence whereby unnatural amino acids can be more efficiently incorporated than original natural amino acids. More specifically, tyrosyl-tRNA synthetases having been modified at the amino acid(s) at the 37- and/or 195-positions. A method of modifying tyrosyl-tRNA synthetase characterized in that the position of an amino acid to be modified is determined based on the 3-D structure of a tyrosyl-AMP and of tyrosyl synthetase complex and an amino acid to which this enzyme binds.

Tyrosyl-tRNA synthetase variants

A novel nucleic acid (aptamer) which binds specifically to the target protein of Ras, more particularly, a novel RNA aptamer which binds specifically to Raf-1; a method for screening an RNA capable of binding specifically to the target protein of Ras which comprises selecting an RNA capable of binding to the target protein of Ras from a pool of RNAs having various base sequences; a method for regulating the signal transduction causing the proliferation or differentiation of cells by using the above-described nucleic acid; and medicinal compositions with the use of the same.

Growing community of inventors

Yokohama, Japan

Kensaku Sakamoto