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    SORTASE MOLECULES AND USES THEREOF
    1.
    发明申请
    SORTASE MOLECULES AND USES THEREOF 审中-公开
    筛选分子及其用途

    公开(公告)号：WO2016014501A1

    公开(公告)日：2016-01-28

    申请号：PCT/US2015/041293

    申请日：2015-07-21

    Applicant: NOVARTIS AG ,  GUIMARAES, Carla

    Inventor： GUIMARAES, Carla

    IPC: C12N9/64

    CPC classification number: C12N9/52 ,  A61K47/65 ,  C12Y304/2207

    Abstract: This application provides mutant sortase molecules and methods of making and using them. In a first aspect, disclosed herein, are sortase molecules having one or a combination of mutations. In an embodiment, a sortase molecule is optimized for a parameter of enzyme performance, e.g., Ca++ dependency (or independency) or reaction rate.

    Abstract translation: 该应用提供突变分选酶分子及其制备和使用方法。 在本文公开的第一方面，是具有一种或一种突变组合的分选酶分子。 在一个实施方案中，分选酶分子针对酶性能的参数（例如Ca ++依赖性（或独立性）或反应速率）进行了优化。

    METHOD FOR SELECTION AND PRODUCTION OF TAILOR-MADE HIGHLY SELECTIVE AND MULTI-SPECIFIC TARGETING ENTITIES CONTAINING AT LEAST TWO DIFFERENT BINDING ENTITIES AND USES THEREOF
    2.
    发明申请
    METHOD FOR SELECTION AND PRODUCTION OF TAILOR-MADE HIGHLY SELECTIVE AND MULTI-SPECIFIC TARGETING ENTITIES CONTAINING AT LEAST TWO DIFFERENT BINDING ENTITIES AND USES THEREOF 审中-公开
    选择和生产包含至少两种不同结合实体的定制高选择性和多特异性靶向实体的方法及其用途

    公开(公告)号：WO2014001324A9

    公开(公告)日：2014-06-19

    申请号：PCT/EP2013063258

    申请日：2013-06-25

    Applicant: HOFFMANN LA ROCHE ,  HOFFMANN LA ROCHE

    Inventor： FENN SEBASTIAN ,  KOPETZKI ERHARD ,  TIEFENTHALER GEORG

    IPC: C07K16/00 ,  C07K16/46 ,  C12N9/16 ,  C12N9/52 ,  C12N9/80 ,  C12P21/06

    CPC classification number: C07K16/32 ,  C07K16/468 ,  C07K2317/24 ,  C07K2317/52 ,  C07K2317/55 ,  C07K2317/64 ,  C12N9/52 ,  C12Y304/2207

    Abstract: Herein is reported a method for producing a bispecific antibody comprising the step of incubating (i) an antibody Fab fragment or a sc Fv antibody comprising within the 20 C-terminal amino acid residues the amino acid sequence LPX1TG (SEQ ID NO: 01), (ii) a one-armed antibody comprising a full length antibody heavy chain, a full length antibody light chain, and an Fc-heavy chain, whereby the full length antibody heavy chain and the full length antibody light chain are cognate antibody chains that thereof forms an antigen binding site, whereby the full length antibody heavy chain and the Fc-heavy chain are covalently linked to each other via one or more disulfide bonds forming an antibody hinge region, and whereby the Fc-heavy chain has an oligoglycine amino acid sequence at its N-terminus, and (iii) a Sortase A enzyme.

    Abstract translation: 本文报道了用于产生双特异性抗体的方法，其包括以下步骤：将（i）在20个C端氨基酸残基内包含氨基酸序列LPX1TG（SEQ ID NO：01）的抗体Fab片段或sc Fv抗体， （ii）包含全长抗体重链，全长抗体轻链和Fc-重链的单臂抗体，其中全长抗体重链和全长抗体轻链是其同源抗体链 形成抗原结合位点，由此全长抗体重链和Fc-重链经由形成抗体铰链区的一个或多个二硫键彼此共价连接，并且其中Fc-重链具有低聚甘氨酸氨基酸序列 在其N-末端，和（iii）分选酶A酶。

    METHOD FOR SELECTION AND PRODUCTION OF TAILOR-MADE HIGHLY SELECTIVE AND MULTI-SPECIFIC TARGETING ENTITIES CONTAINING AT LEAST TWO DIFFERENT BINDING ENTITIES AND USES THEREOF
    3.
    发明申请
    METHOD FOR SELECTION AND PRODUCTION OF TAILOR-MADE HIGHLY SELECTIVE AND MULTI-SPECIFIC TARGETING ENTITIES CONTAINING AT LEAST TWO DIFFERENT BINDING ENTITIES AND USES THEREOF 审中-公开
    选择和生产包含至少两个不同绑定实体的定制选择性和多特定目标实体的方法及其用途

    公开(公告)号：WO2014001324A1

    公开(公告)日：2014-01-03

    申请号：PCT/EP2013/063258

    申请日：2013-06-25

    Applicant: HOFFMANN-LA ROCHE AG ,  HOFFMANN-LA ROCHE INC.

    Inventor： FENN, Sebastian ,  KOPETZKI, Erhard ,  TIEFENTHALER, Georg

    IPC: C07K16/00 ,  C12N9/16 ,  C12N9/52 ,  C12N9/80 ,  C12P21/06 ,  C07K16/46

    CPC classification number: C07K16/32 ,  C07K16/468 ,  C07K2317/24 ,  C07K2317/52 ,  C07K2317/55 ,  C07K2317/64 ,  C12N9/52 ,  C12Y304/2207

    Abstract: Herein is reported a method for producing a bispecific antibody comprising the step of incubating (i) an antibody Fab fragment or a sc Fv antibody comprising within the 20 C-terminal amino acid residues the amino acid sequence LPX1TG (SEQ ID NO: 01), (ii) a one-armed antibody comprising a full length antibody heavy chain, a full length antibody light chain, and an Fc-heavy chain, whereby the full length antibody heavy chain and the full length antibody light chain are cognate antibody chains that thereof forms an antigen binding site, whereby the full length antibody heavy chain and the Fc-heavy chain are covalently linked to each other via one or more disulfide bonds forming an antibody hinge region, and whereby the Fc-heavy chain has an oligoglycine amino acid sequence at its N-terminus, and (iii) a Sortase A enzyme.

    Abstract translation: 本文报道了一种制备双特异性抗体的方法，其包括以下步骤：（i）抗体Fab片段或sc Fv抗体，其包含在20个C末端氨基酸残基内的氨基酸序列LPX1TG（SEQ ID NO：01）， （ii）包含全长抗体重链，全长抗体轻链和Fc重链的单臂抗体，其中全长抗体重链和全长抗体轻链是其中的同源抗体链 形成抗原结合位点，由此全长抗体重链和Fc重链通过形成抗体铰链区的一个或多个二硫键共价连接，由此Fc重链具有寡聚氨基酸序列 在其N-末端，和（iii）分选酶A酶。

    자가 절단 카세트를 포함하는 단백질 정제방법 및 이의 용도
    6.
    发明申请
    자가 절단 카세트를 포함하는 단백질 정제방법 및 이의 용도 审中-公开
    用于精制包含自切割蛋白的蛋白质的方法及其用途

    公开(公告)号：WO2014175690A1

    公开(公告)日：2014-10-30

    申请号：PCT/KR2014/003639

    申请日：2014-04-25

    Applicant: (재) 스크립스코리아 항체연구원

    Inventor： 송병두 ,  윤지선 ,  최효정 ,  김혜인 ,  이응석

    IPC: C07K19/00 ,  C12N15/62 ,  C12N15/70 ,  C07K1/16

    CPC classification number: C07K16/00 ,  C07K1/22 ,  C07K2319/00 ,  C07K2319/21 ,  C07K2319/30 ,  C07K2319/41 ,  C07K2319/42 ,  C07K2319/50 ,  C12N9/50 ,  C12N9/52 ,  C12N15/1093 ,  C12Y304/2207

    Abstract: 본 발명은 아미노 말단에서부터 목적 단백질, LPXTG의 아미노산 서열로 이루어지는 펩타이드, 솔테이즈 A 절단 기능 도메인 및 태그의 순서로 포함하는 자가절단 융합 단백질, 이를 코딩하는 핵산, 본 발명의 핵산을 포함하는 발현벡터, 및 본 발명의 발현벡터로 형질전환된 세포에 관한 것이다. 또한, 본 발명은 상기 형질전환된 세포를 배양하여 용해 및 정제하는 단계를 포함하는 목적 단백질의 정제방법에 관한 것이다. 본 발명은 솔테이즈 A 절단 기능 도메인과 이의 인식 서열인 LPXTG의 아미노산 서열로 구성되는 펩타이드로 이루어진 자가 절단 카세트를 포함하는 자가 절단 융합 단백질에 관한 것으로, 목적 단백질의 정제 및 태그 제거 과정을 별도의 과정이 아닌 한 번의 정제 과정으로 완료할 수 있다는 점에서 매우 유용하다. 특히, 목적 단백질의 위치가 아미노 말단에 존재함으로써 융합 단백질의 컬럼 결합 능력과 자가 절단 능력을 높인 점, 태그가 제거된 목적 단백질을 고순도로 얻을 수 있는 점, 절단-버퍼를 통해 정제 및 태그 제거 과정이 한 번의 과정으로 완료되어 정제에 필요한 시간 및 노력이 획기적으로 감소되는 점, 및 한 번의 단계로 이루어지기 때문에 수득 단백질의 손실이 감소되는 점에서 고순도 및 대량의 단백질이 필요한 다양한 분야에서 널리 이용될 수 있다. 특히, 치료용항체-약물 결합체 제조에 유용하다.

    Abstract translation: 本发明涉及自切割融合蛋白，其包括依次为靶蛋白，由LPXTG的氨基酸序列组成的肽，分选酶A切割功能结构域和来自氨基末端的标签; 其编码核酸; 包含核酸的本发明的表达载体; 和由本发明的表达载体转化的细胞。 本发明还涉及包括对转化细胞进行培养，溶解和精制的步骤的靶蛋白质精制方法。 本发明涉及自切割融合蛋白，其包括自切割盒，其由分选酶A切割功能结构域和由作为其识别序列的LPXTG氨基酸序列组成的肽，并且非常有用，因为 精制目标蛋白质并消除其标签的步骤可以通过单独的精制步骤而不是单独的步骤来实现。 特别地，本发明可用于需要大量高纯度蛋白质的各种领域，因为存在于氨基末端的靶蛋白质可提高融合蛋白，靶蛋白的柱联合能力和自切割能力 以高纯度获得标签的清除，切割缓冲液使得通过单一步骤完成并消除标签的步骤，显着减少了精炼所需的时间和精力，并且通过单步骤的成就减少了 蛋白质产量损失。 特别地，本发明可用于制备治疗性抗体 - 药物复合物。

    METHODS FOR GENERATING ENGINEERED ENZYMES
    8.
    发明申请
    METHODS FOR GENERATING ENGINEERED ENZYMES 审中-公开
    产生工程酶的方法

    公开(公告)号：WO2016061199A3

    公开(公告)日：2016-06-09

    申请号：PCT/US2015055494

    申请日：2015-10-14

    Applicant: RES DEV FOUNDATION

    Inventor： IVERSON BRENT ,  MAREK PETER ,  TAFT JOSEPH

    IPC: C12Q1/37 ,  C12N15/10 ,  C12N15/81 ,  C12Q1/48

    CPC classification number: C12P21/005 ,  C12Q1/37 ,  C12Q1/485 ,  C12Q1/6816 ,  C12Q1/6869 ,  C12Y207/10001 ,  C12Y304/2207 ,  G01N33/5008 ,  G01N2333/91205 ,  G01N2333/96466 ,  G06F19/22

    Abstract: Provided are improved methods for identifying the substrate recognition specificity or activity of a protease, convertase (sortase), or kinase. In some embodiments, methods are provided for identifying the endogenous protease or convertase cleaving patterns (e.g., "cleaveOme") inside the secretory pathway of a living cell. Select embodiments involve aspects of yeast endoplasmic reticulum sequestration screening and next generation sequencing. Methods of producing polypeptides in Kex2 knockout yeast are also provided.

    Abstract translation: 提供了用于鉴定蛋白酶，转化酶（分选酶）或激酶的底物识别特异性或活性的改进方法。 在一些实施方案中，提供了用于鉴定活细胞分泌途径内的内源性蛋白酶或转化酶切割模式（例如，“切割蛋白”）的方法。 选择实施方案涉及酵母内质网隔离筛选和下一代测序的方面。 还提供了在Kex2敲除酵母中产生多肽的方法。

    METHODS FOR GENERATING ENGINEERED ENZYMES
    9.
    发明申请
    METHODS FOR GENERATING ENGINEERED ENZYMES 审中-公开
    生成工程化酶的方法

    公开(公告)号：WO2016061199A2

    公开(公告)日：2016-04-21

    申请号：PCT/US2015/055494

    申请日：2015-10-14

    Applicant: RESEARCH DEVELOPMENT FOUNDATION

    Inventor： IVERSON, Brent ,  MAREK, Peter ,  TAFT, Joseph

    IPC: C12Q1/37

    CPC classification number: C12P21/005 ,  C12Q1/37 ,  C12Q1/485 ,  C12Q1/6816 ,  C12Q1/6869 ,  C12Y207/10001 ,  C12Y304/2207 ,  G01N33/5008 ,  G01N2333/91205 ,  G01N2333/96466 ,  G06F19/22

    Abstract: Provided are improved methods for identifying the substrate recognition specificity or activity of a protease, convertase (sortase), or kinase. In some embodiments, methods are provided for identifying the endogenous protease or convertase cleaving patterns (e.g., "cleaveOme") inside the secretory pathway of a living cell. Select embodiments involve aspects of yeast endoplasmic reticulum sequestration screening and next generation sequencing. Methods of producing polypeptides in Kex2 knockout yeast are also provided.

    Abstract translation: 提供了用于鉴定蛋白酶，转化酶（分选酶）或激酶的底物识别特异性或活性的改进方法。 在一些实施方案中，提供了用于鉴定活细胞分泌途径内的内源性蛋白酶或转化酶切割模式（例如“切割蛋白”）的方法。 选择实施方案涉及酵母内质网螯合筛选和下一代测序的方面。 还提供了在Kex2敲除酵母中生产多肽的方法。

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