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    • 5. 发明申请
    • METHODS FOR THE IDENTIFICATION OF PI3K INTERACTING MOLECULES AND FOR THE PURIFICATION OF PI3K
    • PI3K相互作用分子鉴定方法和PI3K纯化方法
    • WO2008015013A1
    • 2008-02-07
    • PCT/EP2007/006887
    • 2007-08-03
    • CELLZOME AGBERGAMINI MOORE, GiovannaCANSFIELD, AndrewRAMSDEN, NigelNEUBAUER, Gitte
    • BERGAMINI MOORE, GiovannaCANSFIELD, AndrewRAMSDEN, NigelNEUBAUER, Gitte
    • G01N33/573C12Q1/48
    • C12Q1/485G01N2500/02
    • The present invention relates to a method for the identification of a PI3K interacting compound, comprising the steps of a) providing a protein preparation containing PI3K, b) contacting the protein preparation with phenylthiazole ligand 1 immobilized on a solid support under conditions allowing the formation of a phenylthiazole ligand 1 - PI3K complex, c) incubating the phenylthiazole ligand 1 - PI3K complex with a given compound, and d) determining whether the compound is able to separate PI3K from the immobilized phenylthiazole ligand 1. Furthermore, the present invention relates to a method for the identification of a PI3K interacting compound, comprising the steps of a) providing a protein preparation containing PI3K, b) contacting the protein preparation with phenylthiazole ligand 1 immobilized on a solid support and with a given compound under conditions allowing the formation of a phenylthiazole ligand 1 - PI3K complex, and c) detecting the phenylthiazole ligand 1 - PI3K complex formed in step b). Furthermore, the present invention relates to a method for the identification of a PI3 K interacting compound, comprising the steps of a) providing two aliquots of a protein preparation containing PI3K, b) contacting one aliquot with the phenylthiazole ligand 1 immobilized on a solid support under conditions allowing the formation of a phenylthiazole ligand 1 - PI3K complex, c) contacting the other aliquot with the phenylthiazole ligand 1 immobilized on a solid support and with a given compound under conditions allowing the formation of a phenylthiazole ligand 1 - PI3K complex, and d) determining the amount of the phenylthiazole ligand 1 - PI3K complex formed in steps b) and c). Furthermore, the present invention realtes to a method for the identification of a PI3K interacting compound, comprising the steps of a) providing two aliquots comprising each at least one cell containing PI3K, b) incubating one aliquot with a given compound, c) harvesting the cells of each aliquot, d) lysing the cells in order to obtain protein preparations, e) contacting the protein preparations with the phenylthiazole ligand 1 immobilized on a solid support under conditions allowing the formation of a phenylthiazole ligand 1 - PI3K complex, and f) determining the amount of the phenylthiazole ligand 1 - PI3K complex formed in each aliquot in step e).
    • 本发明涉及一种鉴定PI3K相互作用化合物的方法,包括以下步骤:a)提供含有PI3K的蛋白质制剂,b)使蛋白制备物与固定在固相载体上的苯基噻唑配体1接触, 苯基噻唑配体1-PI3K复合物,c)将苯基噻唑配体1-PI3K复合物与给定化合物一起孵育,以及d)测定该化合物是否能够从固定的苯基噻唑配体1分离PI3K。此外,本发明涉及 用于鉴定PI3K相互作用化合物的方法,包括以下步骤:a)提供含有PI3K的蛋白质制剂,b)使所述蛋白质制剂与固定在固体支持物上的苯基噻唑配体1和给定化合物在允许形成 苯基噻唑配体1-PI3K复合物,和c)检测苯基噻唑配体1-PI3K互补 x在步骤b)中形成。 此外,本发明涉及用于鉴定PI3K相互作用化合物的方法,包括以下步骤:a)提供含有PI3K的蛋白质制剂的两个等分试样,b)使一个等分试样与固定在固体支持物上的苯基噻唑配体1接触 在允许形成苯基噻唑配体1-PI3K复合物的条件下,c)使其他等分试样与固定在固体载体上的苯基噻唑配体1和允许形成苯基噻唑配体1-PI3K复合物的条件下与给定化合物接触,以及 d)测定步骤b)和c)中形成的苯基噻唑配体1-PI3K复合物的量。 此外,本发明涉及用于鉴定PI3K相互作用化合物的方法,其包括以下步骤:a)提供两个等分试样,其包含每个至少一个含有PI3K的细胞,b)将一个等分试样与给定化合物一起孵育,c) 每个等分试样的细胞,d)裂解细胞以获得蛋白质制剂,e)在允许形成苯基噻唑配体1-PI3K复合物的条件下,使蛋白质制剂与固定在固体支持物上的苯基噻唑配体1接触,以及f) 测定在步骤e)中在每个等分试样中形成的苯基噻唑配体1-PI3K复合物的量。