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    • 8. 发明授权
    • E2 displacement assay for identifying inhibitors of HPV
    • 用于鉴定HPV抑制剂的E2位移测定
    • US06825176B2
    • 2004-11-30
    • US10358790
    • 2003-02-05
    • Peter WhiteChristiane Yoakim
    • Peter WhiteChristiane Yoakim
    • A01N4304
    • C07D407/12C07D491/10G01N33/532G01N33/533G01N33/534G01N33/535G01N33/542G01N33/56983G01N33/581G01N33/582G01N33/583G01N33/60G01N2333/025G01N2500/10
    • The present invention generally relates to an assay for identifying inhibitors of Human Papillomavirus (HPV), comprising: a) contacting a HPV E2 transactivation domain with a probe to form a E2:probe complex and measuring a signal from said probe to establish a base line level; b) incubating the E2:probe complex with a test compound and measuring the signal from said probe; c) comparing the signal from step b) with the signal from step a); wherein said probe is a compound of formula (I) or its enantiomers or diastereoisomers thereof: wherein R1, A, X, W, Y, R3 and R4 are as defined herein; or a derivative thereof, wherein said derivative is a probe of formula (I) labeled with a detectable label or an affinity tag, wherein wavy lines represent bonds of unspecified stereochemistry; and wherein said signal is selected from: fluorescence, resonance energy transfer, time resolved fluorescence, radioactivity, fluorescence polarization, change in the intrinsic spectral properties, luminescence and plasma-resonance; whereby a modulation in said signal is an indication that said test compound binds to said transactivation domain.
    • 本发明一般涉及用于鉴定人乳头瘤病毒(HPV)抑制剂的测定法,其包括:a)使HPV E2反式激活结构域与探针接触以形成E2:探针复合物并测量来自所述探针的信号以建立基线 b)将E2:探针复合物与测试化合物孵育并测量来自所述探针的信号; c)将来自步骤b)的信号与来自步骤a)的信号进行比较;其中所述探针是式(I)的化合物, 或其对映体或非对映异构体:其中R 1,A,X,W,Y,R 3和R 4如本文所定义; 或其衍生物,其中所述衍生物是用可检测标记或亲和标签标记的式(I)的探针,其中波浪线表示未指定的立体化学键; 并且其中所述信号选自:荧光,共振能量转移,时间分辨荧光,放射性,荧光偏振,固有光谱性质的变化,发光和等离子体共振; 由此所述信号中的调节是所述测试化合物结合所述反式激活结构域的指示。
    • 9. 发明授权
    • Method of identifying potential inhibitors of human papillomavirus protein E2 using x-ray atomic coordinates
    • 使用x射线原子坐标识别人乳头瘤病毒蛋白E2的潜在抑制剂的方法
    • US07167801B2
    • 2007-01-23
    • US10193460
    • 2002-07-11
    • Dale R. CameronJacques ArchambaultChristiane YoakimPeter WhiteYong Wang
    • Dale R. CameronJacques ArchambaultChristiane YoakimPeter WhiteYong Wang
    • G06F7/58
    • C07D405/12C07D417/12C07K14/005C07K2299/00C12N2710/20022
    • A crystallizable composition, comprising an human papillomavirus (HPV-11) E2 transactivation domain (TAD)-like polypeptide of SEQ ID NO. 2 complexed with an inhibitor L (sodium (2R,3R,4S,5R)-5-(3,4-dichlorophenyl)-5′-methyl-1′,3′-dioxo-4-({[4-(1,2,3-thiadiazol-4-yl)phenyl]amino}carbonyl)-1′,3′,4,5-tetrahydro-3H-spiro[furan-2,2′-indene]-3-carboxylate). The invention also provides a method for producing the crystallized HPV E2 TAD-inhibitor complex (HPV E2 TAD-L) comprising: a) mixing purified HPV E2 TAD, contained in a purification buffer, with solublized inhibitor L to generate a complex solution containing the HPV E2 TAD-L complex; and b) crystallizing the complex from a) in a crystallization buffer. The invention also provides a method for producing crystallized apo HPV E2 TAD, comprising: a) mixing apo HPV E2 TAD, contained in a purification buffer, with a crystallization buffer.X-ray crystal structure coordinates the HPV E2 TAD-L complex, are also provided, which define an inhibitor binding pocket. The inhibitor binding pocket is useful for screening potential small molecule inhibitors that bind to the pocket that may be inhibitors of papillomavirus infection.
    • 一种可结晶组合物,其包含SEQ ID NO.1的人乳头状瘤病毒(HPV-11)E2反式激活结构域(TAD)样多肽。 2与抑制剂L((2R,3R,4S,5R)-5-(3,4-二氯苯基)-5'-甲基-1',3'-二氧代-4 - ({[4-(1 ,2,3-噻二唑-4-基)苯基]氨基}羰基)-1',3',4,5-四氢-3H-螺[呋喃-2,2'-茚] -3-羧酸乙酯)。 本发明还提供了一种生产结晶的HPV E2 TAD抑制剂复合物(HPV E2 TAD-L)的方法,包括:a)将纯化缓冲液中纯化的HPV E2 TAD与溶出的抑制剂L混合以产生含有 HPV E2 TAD-L复合物; 和b)从结晶缓冲液中的a)结晶复合物。 本发明还提供了一种用于生产结晶apo HPV E2 TAD的方法,其包括:a)将包含在纯化缓冲液中的apo HPV E2 TAD与结晶缓冲液混合。 还提供X射线晶体结构协调HPV E2 TAD-L复合物,其定义了抑制剂结合口袋。 抑制剂结合口袋可用于筛选结合口袋的可能是乳头瘤病毒感染抑制剂的潜在小分子抑制剂。