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    tRNA SYNTHESIS METHOD, NUCLEIC ACID, AMINOACYL tRNA SYNTHESIS METHOD, AND PROCESS FOR PRODUCTION OF PROTEIN INTRODUCED WITH UNNATURAL AMINO ACID
    1.
    发明申请
    tRNA SYNTHESIS METHOD, NUCLEIC ACID, AMINOACYL tRNA SYNTHESIS METHOD, AND PROCESS FOR PRODUCTION OF PROTEIN INTRODUCED WITH UNNATURAL AMINO ACID 审中-公开
    tRNA合成方法，核酸，氨基酸tRNA合成方法和生产引发氨基酸的蛋白质的方法

    公开(公告)号：US20100304431A1

    公开(公告)日：2010-12-02

    申请号：US11997528

    申请日：2006-08-02

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takatsugu Kobayashi

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takatsugu Kobayashi

    IPC分类号： C12P21/02 ,  C12P19/34

    CPC分类号： C12N9/93 ,  C12N15/67 ,  C12P19/34 ,  C12P21/02

    摘要： The present invention relates to a process for producing a protein having an unnatural amino acid introduced therein, the process including: expressing in a eukaryotic cell an aminoacyl-tRNA synthetase, a nucleic acid having a sequence containing a eukaryote-derived tRNA nucleotide sequence linked to the 5′ end of a tRNA nucleotide sequence that is ligated with to an unnatural amino acid in the presence of the aminoacyl-tRNA synthetase, an unnatural amino acid, and a gene of a desired protein having a nonsense mutation at a predetermined position, to integrate the unnatural amino acid at the nonsense mutation position into the protein, thereby expressing a protein having an unnatural amino acid introduced therein.

    摘要翻译： 本发明涉及一种生产引入其中具有非天然氨基酸的蛋白质的方法，所述方法包括：在真核细胞中表达氨基-tRNA合成酶，具有包含与真核生物衍生的tRNA核苷酸序列连锁的序列的核酸 在氨基-tRNA合成酶，非天然氨基酸和在预定位置具有无义突变的所需蛋白质的基因，在预定位置具有无义突变的基因，与非天然氨基酸连接的tRNA核苷酸序列的5'末端， 将无义突变位置处的非天然氨基酸整合到蛋白质中，从而表达引入其中的非天然氨基酸的蛋白质。

    Process for production of non-natural protein having ester bond therein
    2.
    发明授权
    Process for production of non-natural protein having ester bond therein 失效
    生产其中具有酯键的非天然蛋白质的方法

    公开(公告)号：US08785152B2

    公开(公告)日：2014-07-22

    申请号：US12744322

    申请日：2008-11-21

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takahito Mukai ,  Takatsugu Kobayashi

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takahito Mukai ,  Takatsugu Kobayashi

    IPC分类号： C12P21/00 ,  C12N9/00 ,  C12N15/52 ,  C12N1/21

    CPC分类号： C12N15/67 ,  C12N9/93 ,  C12P21/02

    摘要： 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 α-hydroxy acid: (a) an aminoacyl-tRNA synthetase which can activate the α-hydroxy acid; (b) suppressor tRNA which can bind to the α-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.

    摘要翻译： 在其多肽主链中具有至少一个酯键的非天然蛋白质通过使用核糖体中的体内翻译系统来合成。 以下组分（a）至（c）在α-羟基酸存在下在细胞或细胞提取溶液中表达：（a）可活化α-羟基酸的氨基酰-tRNA合成酶; （b）在氨基-tRNA合成酶存在下可结合α-羟基酸的抑制tRNA; 和（c）编码期望蛋白质的基因，所述蛋白质在期望的位点具有无义突变或帧位移突变。

    Mutant pyrrolysyl-tRNA synthetase, and method for production of protein having non-natural amino acid integrated therein by using the same
    3.
    发明授权
    Mutant pyrrolysyl-tRNA synthetase, and method for production of protein having non-natural amino acid integrated therein by using the same 有权
    突变型吡咯赖氨酰-tRNA合成酶，以及通过使用它们生成其中含有非天然氨基酸的蛋白质的方法

    公开(公告)号：US08735093B2

    公开(公告)日：2014-05-27

    申请号：US12727037

    申请日：2010-03-18

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takatsugu Kobayashi

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takatsugu Kobayashi

    IPC分类号： C12P21/00 ,  C12N9/00 ,  C12N15/52 ,  C12N15/63

    CPC分类号： C12N9/93 ,  C12P21/00 ,  C12P21/02

    摘要： Method for incorporating a lysine derivative (particularly an Nε-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.

    摘要翻译： 引入适合作为非天然的有重要原子，硒，反应性官能团，荧光基团或交联剂等有用官能团的赖氨酸衍生物（特别是N，N-苄氧羰基赖氨酸（Z-Lys）衍生物）的方法 氨基酸，以位点特异性方式转化成所需的蛋白质。 突变型吡咯赖氨酰-tRNA合成酶具有至少一个氨基酸残基的取代，该氨基酸残基选自306位的酪氨酸残基，309位的亮氨酸残基和348位的半胱氨酸残基，每个构成吡咯赖氨酰-tRNA的氨基酸序列中的吡咯赖氨酸结合位点 SEQ ID NO：2的合成酶。 氨基酸残基的取代是：位置306处的甘氨酸或丙氨酸残基的酪氨酸残基，309位的亮氨酸残基被甘氨酸或丙氨酸残基取代，和/或348位的半胱氨酸残基被缬氨酸，丝氨酸或丙氨酸残基取代 。

    PROCESS FOR PRODUCTION OF NON-NATURAL PROTEIN HAVING ESTER BOND THEREIN
    4.
    发明申请
    PROCESS FOR PRODUCTION OF NON-NATURAL PROTEIN HAVING ESTER BOND THEREIN 失效
    生产非蛋白质的非天然蛋白质的方法

    公开(公告)号：US20110136168A1

    公开(公告)日：2011-06-09

    申请号：US12744322

    申请日：2008-11-21

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takahito Mukai ,  Takatsugu Kobayashi

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Tatsuo Yanagisawa ,  Takahito Mukai ,  Takatsugu Kobayashi

    IPC分类号： C12P21/02

    CPC分类号： C12N15/67 ,  C12N9/93 ,  C12P21/02

    摘要： 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 α-hydroxy acid: (a) an aminoacyl-tRNA synthetase which can activate the α-hydroxy acid; (b) suppressor tRNA which can bind to the α-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.

    摘要翻译： 在其多肽主链中具有至少一个酯键的非天然蛋白质通过使用核糖体中的体内翻译系统来合成。 以下组分（a）至（c）在α-羟基酸存在下在细胞或细胞提取溶液中表达：（a）可活化α-羟基酸的氨基酰-tRNA合成酶; （b）在氨基-tRNA合成酶存在下可结合α-羟基酸的抑制tRNA; 和（c）编码期望蛋白质的基因，所述蛋白质在期望的位点具有无义突变或帧位移突变。

    Polypeptide having activity of aminoacyl-tRNA synthetase and use thereof
    9.
    发明申请
    Polypeptide having activity of aminoacyl-tRNA synthetase and use thereof 审中-公开
    具有氨基-tRNA合成酶活性的多肽及其用途

    公开(公告)号：US20090226966A1

    公开(公告)日：2009-09-10

    申请号：US12322782

    申请日：2009-02-05

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Kenji Oki ,  Takahito Mukai

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Kenji Oki ,  Takahito Mukai

    IPC分类号： C12N9/00 ,  C12N15/52 ,  C12P21/02

    CPC分类号： C12N9/93

    摘要： 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.

    摘要翻译： 根据本发明的多肽包括：通过改变ArgRS，CysRS，MetRS，GlnRS，GluRS，LysRS，TyrRS或TrpRS获得的改变的多肽，使得识别出非天然氨基酸; 和衍生自PheRS，LeuRS，IleRS，ValRS，AlaRS，ProRS或ThrRS的编辑多肽，所述编辑多肽已插入Rossman折叠N结构域和Rossman折叠C结构域之间， 存在于改变的多肽中，或者与改变的多肽的N末端结合。 因此提供了对非天然氨基酸表现出高底物特异性的新aaRS和涉及使用这种aaRS的技术。

    Method for producing proteins comprising non-natural amino acids incorporated therein
    10.
    发明授权
    Method for producing proteins comprising non-natural amino acids incorporated therein 失效
    包含非天然氨基酸的蛋白质的生产方法

    公开(公告)号：US08642291B2

    公开(公告)日：2014-02-04

    申请号：US13470752

    申请日：2012-05-14

    申请人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Fumie Iraha

    发明人： Shigeyuki Yokoyama ,  Kensaku Sakamoto ,  Fumie Iraha

    IPC分类号： C12N9/00 ,  A61K45/06 ,  C12P21/02 ,  A61K38/00

    CPC分类号： C12P21/02 ,  C12N9/93 ,  C12N2510/02

    摘要： 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.

    摘要翻译： 产生含有非天然氨基酸的蛋白质可以涉及将基因导入真核细胞型细胞内并敲入固有基因。 要引入的基因包括具有对非天然氨基酸的特异性增强的真核生物型氨基酰基tRNA合成酶突变体的基因，与对相似天然氨基酸的特异性相比，以及能够结合非天然氨基酸的非天然氨基酸的tRNA基因 在真核生物型氨酰tRNA合成酶突变体存在下的氨基酸。 要敲除的固有基因包括编码具有对天然氨基酸具有特异性的氨基酰基tRNA合成酶的基因和能够在固有氨基tRNA合成酶存在下与天然氨基酸结合的tRNA基因。