公开(公告)号：US07592140B2

公开(公告)日：2009-09-22

申请号：US11078951

申请日：2005-03-11

申请人： Kazuishi Kubota ,  Kaori Nakahara ,  Ayako Hara ,  Yohei Ozeki ,  Yasuteru Iijima

发明人： Kazuishi Kubota ,  Kaori Nakahara ,  Ayako Hara ,  Yohei Ozeki ,  Yasuteru Iijima

IPC分类号： C12N15/55 ,  C12Q1/68 ,  C12N9/16

CPC分类号： C12Q1/44 ,  A61K38/00 ,  C12N9/22 ,  G01N2500/00

摘要： Polypeptides having 2′,5′-oligoadenylate phosphodiesterase activity, antibodies against the polypeptides, polynucleotides coding for the polypeptides, recombinant plasmid DNAs into which the polynucleotides have been inserted, host cells transformed by the recombinant plasmid DNAs, processes for screening for 2′,5′-oligoadenylate phosphodiesterase activity inhibitor substances, and processes for screening for 2′,5′-oligoadenylate phosphodiesterase expression regulation inhibitor substances. The polypeptides, polynucleotides and host cells are useful for searching for therapeutic agents for viral infections and tumors.

摘要翻译： 具有2'，5'-寡腺苷酸磷酸二酯酶活性的多肽，针对多肽的抗体，编码多肽的多核苷酸，插入多核苷酸的重组质粒DNA，由重组质粒DNA转化的宿主细胞，筛选2'， 5'-寡腺苷酸磷酸二酯酶活性抑制剂物质，以及筛选2'，5'-寡腺苷酸磷酸二酯酶表达调节抑制物质的方法。 多肽，多核苷酸和宿主细胞可用于寻找病毒感染和肿瘤的治疗剂。

公开(公告)号：US20050272119A1

公开(公告)日：2005-12-08

申请号：US11078951

申请日：2005-03-11

申请人： Kazuishi Kubota ,  Kaori Nakahara ,  Ayako Hara ,  Yohei Ozeki ,  Yasuteru Iijima

发明人： Kazuishi Kubota ,  Kaori Nakahara ,  Ayako Hara ,  Yohei Ozeki ,  Yasuteru Iijima

IPC分类号： A61K38/00 ,  A61P31/12 ,  A61P35/00 ,  C07K16/40 ,  C12N1/21 ,  C12N9/22 ,  C12Q1/44 ,  C12N9/16 ,  C07H21/04

CPC分类号： C12Q1/44 ,  A61K38/00 ,  C12N9/22 ,  G01N2500/00

摘要： Polypeptides having 2′,5′-oligoadenylate phosphodiesterase activity, antibodies against the polypeptides, polynucleotides coding for the polypeptides, recombinant plasmid DNAs into which the polynucleotides have been inserted, host cells transformed by the recombinant plasmid DNAs, processes for screening for 2′,5′-oligoadenylate phosphodiesterase activity inhibitor substances, and processes for screening for 2′,5′-oligoadenylate phosphodiesterase expression regulation inhibitor substances. The polypeptides, polynucleotides and host cells are useful for searching for therapeutic agents for viral infections and tumors.

摘要翻译： 具有2'，5'-寡腺苷酸磷酸二酯酶活性的多肽，针对多肽的抗体，编码多肽的多核苷酸，插入多核苷酸的重组质粒DNA，由重组质粒DNA转化的宿主细胞，筛选2'， 5'-寡腺苷酸磷酸二酯酶活性抑制剂物质，以及筛选2'，5'-寡腺苷酸磷酸二酯酶表达调节抑制物质的方法。 多肽，多核苷酸和宿主细胞可用于寻找病毒感染和肿瘤的治疗剂。

公开(公告)号：US20100248227A1

公开(公告)日：2010-09-30

申请号：US12376434

申请日：2007-08-03

申请人： Futoshi Nara ,  Kiyoaki Yonesu ,  Kazuishi Kubota

发明人： Futoshi Nara ,  Kiyoaki Yonesu ,  Kazuishi Kubota

IPC分类号： C12Q1/68 ,  G01N33/573 ,  C12P7/62

CPC分类号： C12Q1/485 ,  C12N9/1205 ,  C12P17/00 ,  G01N2500/00

摘要： The objects of the present invention are: elucidation of an enzyme that phosphorylates in vivo a compound such as (2R)-2-amino-2-methyl-4-[5-(5-phenylpentanoyl)thiophen-2-yl]butan-1-ol and provision of a method of phosphorylating the aforementioned compound; provision of a method of screening for a substance phosphorylated by the aforementioned enzyme; provision of a method of determining the ability of a subject to phosphorylate a test compound.Provision of a method of phosphorylating the aforementioned compound using a human fructosamine-3-kinase-related protein and/or fructosamine-3-kinase, and a method of determining the ability of a subject to phosphorylate a test compound.

摘要翻译： 本发明的目的是：阐明在体内磷酸化化合物如（2R）-2-氨基-2-甲基-4- [5-（5-苯基戊酰基）噻吩-2-基]丁-1-醇的化合物的酶， 提供磷酸化上述化合物的方法; 提供筛选由上述酶磷酸化的物质的方法; 提供确定受试者磷酸化测试化合物的能力的方法。 提供使用人果糖胺-3-激酶相关蛋白和/或果糖胺-3-激酶磷酸化上述化合物的方法，以及确定受试者磷酸化试验化合物的能力的方法。

公开(公告)号：US08071300B2

公开(公告)日：2011-12-06

申请号：US12376434

申请日：2007-08-03

申请人： Futoshi Nara ,  Kiyoaki Yonesu ,  Kazuishi Kubota

发明人： Futoshi Nara ,  Kiyoaki Yonesu ,  Kazuishi Kubota

IPC分类号： C12Q1/68 ,  C12P7/62 ,  G01N33/53

CPC分类号： C12Q1/485 ,  C12N9/1205 ,  C12P17/00 ,  G01N2500/00

摘要： The objects of the present invention are: elucidation of an enzyme that phosphorylates in vivo a compound such as (2R)-2-amino-2-methyl-4-[5-(5-phenylpentanoyl)thiophen-2-yl]butan-1-ol and provision of a method of phosphorylating the aforementioned compound; provision of a method of screening for a substance phosphorylated by the aforementioned enzyme; provision of a method of determining the ability of a subject to phosphorylate a test compound.Provision of a method of phosphorylating the aforementioned compound using a human fructosamine-3-kinase-related protein and/or fructosamine-3-kinase, and a method of determining the ability of a subject to phosphorylate a test compound.

摘要翻译： 本发明的目的是：阐明在体内磷酸化化合物如（2R）-2-氨基-2-甲基-4- [5-（5-苯基戊酰基）噻吩-2-基]丁-1-醇的化合物的酶， 提供磷酸化上述化合物的方法; 提供筛选由上述酶磷酸化的物质的方法; 提供确定受试者磷酸化测试化合物的能力的方法。 提供使用人果糖胺-3-激酶相关蛋白和/或果糖胺-3-激酶磷酸化上述化合物的方法，以及确定受试者磷酸化试验化合物的能力的方法。

公开(公告)号：US20110269161A1

公开(公告)日：2011-11-03

申请号：US13078203

申请日：2011-04-01

申请人： Steven P. Gygi ,  Kazuishi Kubota ,  Judit Villen ,  Yonghao Yu

发明人： Steven P. Gygi ,  Kazuishi Kubota ,  Judit Villen ,  Yonghao Yu

IPC分类号： C12Q1/48 ,  C07K7/08

CPC分类号： C12Q1/485 ,  G01N33/6842 ,  G01N33/6848

摘要： A mass-spectrometry-based method and substrates are provided herein for large scale kinome activity profiling directly from crude lysates using 90 chemically synthesized peptide substrates with amino acid sequences derived from known phosphoproteins. Quantification of peptide phosphorylation rates was achieved via the use of stable isotope labeled synthetic peptides. Half of these peptides immediately or rapidly showed robust and site-specific phosphorylation after incubation with serum-starved HEK293 cell lysate. A method and substrates for obtaining 90 simultaneous activity measurements in a single-reaction format were developed and validated. Activating kinase pathways through insulin or EGF stimulation reproducibly altered the phosphorylation rates of peptides derived from known pathway protein substrates. While examining cell-cycle-specific activities with the panel, a peptide derived from phosphoinositide 3-kinase regulatory subunit demonstrated mitotic and tyrosine-specific phosphorylation, which was confirmed to be a Src kinase site in vivo. The kinome activity profiling strategy was successfully applied with lysates of each of: cells manipulated by various combination of mitogen stimulation, pharmacological perturbation and siRNA-directed kinase knockdown; seven different breast cancer cell lines treated with gefitinib; and each of normal and cancerous tissue samples from renal cell carcinoma patients. This method concurrently measures multiple peptide phosphorylation rates to provide a diagnostic fingerprint pattern for activated kinases, protein phosphatases, modulators of these enzymes, and pathways (kinome) from as little starting material as a few cells.

摘要翻译： 本文提供基于质谱法的方法和底物，用于使用90个具有源自已知磷蛋白的氨基酸序列的化学合成的肽底物直接从粗裂解物进行大规模活性物质分析。 通过使用稳定同位素标记的合成肽来实现肽磷酸化速率的定量。 与血清饥饿的HEK293细胞裂解物孵育后，这些肽中的一半立即或快速显示出强壮的和位点特异性磷酸化。 开发并验证了用于以单反应形式获得90次同时活动测量的方法和底物。 通过胰岛素或EGF刺激激活激酶途径可重复地改变衍生自已知途径蛋白底物的肽的磷酸化速率。 在使用小组检查细胞周期特异性活性的同时，衍生自磷酸肌醇3-激酶调节亚基的肽表现出有丝分裂和酪氨酸特异性磷酸化，这被证实是体内的Src激酶位点。 激酶活性分析策略成功应用于以下各项的裂解物：由促分裂原刺激，药理学扰动和siRNA定向激酶敲低的各种组合操纵的细胞; 用吉非替尼治疗的七种不同的乳腺癌细胞系; 以及来自肾细胞癌患者的正常和癌组织样本。 该方法同时测量多肽磷酸化速率，以提供活化激酶，蛋白磷酸酶，这些酶的调节剂的诊断指纹图谱，以及与几个细胞一样少的起始材料的途径（kinome）。