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    Process for production of non-natural protein having ester bond therein
    5.
    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。

    PROCESS FOR PRODUCTION OF NON-NATURAL PROTEIN HAVING ESTER BOND THEREIN
    6.
    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
    7.
    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。

    POLYPEPTIDE HAVING ACTIVITY OF AMINOACYL-tRNA SYNTHETASE AND USE THEREOF
    8.
    发明申请
    POLYPEPTIDE HAVING ACTIVITY OF AMINOACYL-tRNA SYNTHETASE AND USE THEREOF 有权
    具有氨基酸-TRNA合成酶活性的多肽及其用途

    公开(公告)号：US20120028330A1

    公开(公告)日：2012-02-02

    申请号：US13180209

    申请日：2011-07-11

    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 ,  C07H21/04

    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 constructing recombinant bacterium for producing non-native protein, and utilization of same
    9.
    发明授权
    Method for constructing recombinant bacterium for producing non-native protein, and utilization of same 有权
    构建非天然蛋白质生产重组细菌的方法及其利用方法

    公开(公告)号：US09340790B2

    公开(公告)日：2016-05-17

    申请号：US13704391

    申请日：2011-06-16

    申请人： Shigeyuki Yokoyama ,  Takahito Mukai ,  Kensaku Sakamoto ,  Akiko Matsumoto

    发明人： Shigeyuki Yokoyama ,  Takahito Mukai ,  Kensaku Sakamoto ,  Akiko Matsumoto

    IPC分类号： C12N15/64 ,  C12N15/70 ,  C12N15/67 ,  C12P21/02 ,  C07K14/245 ,  C12N9/00 ,  C12N15/11

    CPC分类号： C12N15/70 ,  C07K14/245 ,  C12N9/93 ,  C12N15/11 ,  C12N15/67 ,  C12P21/02 ,  Y02P20/52

    摘要： 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 α-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.

    摘要翻译： 本发明提供了一种生产非天然蛋白质的重组细菌的新方法，包括：（1）在tRNA识别UAG密码子的细菌中表达tRNA; （2）在细菌中表达氨酰-tRNA合成酶，该氨基-tRNA合成酶用非天然氨基酸或α-羟基酸酰化tRNA; （3）（i）将DNA构建体引入细菌中，所述DNA构建体用于在不存在用于在UAG密码子下终止翻译的释放因子的情况下表达，所述DNA构建体用于表达至少一种选自下组的基因的功能： 当编码释放因子的基因有缺陷和/或引入细菌染色体中的所述至少一个基因的改变时，其失去功能，所述改变用于在所述细菌的染色体中表达所述至少一种基因的功能 不存在释放因子; 和（4）导致编码细菌释放因子的基因是有缺陷的。

    METHOD FOR CONSTRUCTING RECOMBINANT BACTERIUM FOR PRODUCING NON-NATIVE PROTEIN, AND UTILIZATION OF SAME
    10.
    发明申请
    METHOD FOR CONSTRUCTING RECOMBINANT BACTERIUM FOR PRODUCING NON-NATIVE PROTEIN, AND UTILIZATION OF SAME 有权
    用于构建用于生产非蛋白质的重组细菌的方法及其利用

    公开(公告)号：US20130095524A1

    公开(公告)日：2013-04-18

    申请号：US13704391

    申请日：2011-06-16

    申请人： Shigeyuki Yokoyama ,  Takahito Mukai ,  Kensaku Sakamoto ,  Akiko Matsumoto

    发明人： Shigeyuki Yokoyama ,  Takahito Mukai ,  Kensaku Sakamoto ,  Akiko Matsumoto

    IPC分类号： C12N15/70

    CPC分类号： C12N15/70 ,  C07K14/245 ,  C12N9/93 ,  C12N15/11 ,  C12N15/67 ,  C12P21/02 ,  Y02P20/52

    摘要： 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 α-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.

    摘要翻译： 本发明提供了一种生产非天然蛋白质的重组细菌的新方法，包括：（1）在tRNA识别UAG密码子的细菌中表达tRNA; （2）在细菌中表达氨酰-tRNA合成酶，该氨基-tRNA合成酶用非天然氨基酸或α-羟基酸酰化tRNA; （3）（i）将DNA构建体引入细菌中，所述DNA构建体用于在不存在用于在UAG密码子下终止翻译的释放因子的情况下表达，所述DNA构建体用于表达至少一种选自下组的基因的功能： 当编码释放因子的基因有缺陷和/或引入细菌染色体中的所述至少一个基因的改变时，其失去功能，所述改变用于在所述细菌的染色体中表达所述至少一种基因的功能 不存在释放因子; 和（4）导致编码细菌释放因子的基因是有缺陷的。