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首页> 外文期刊>EMBO Journal >SUBSTRATE SPECIFICITY OF THE CDK-ACTIVATING KINASE (CAK) IS ALTERED UPON ASSOCIATION WITH TFIIH
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SUBSTRATE SPECIFICITY OF THE CDK-ACTIVATING KINASE (CAK) IS ALTERED UPON ASSOCIATION WITH TFIIH

机译:CDK-ACTIVATING激酶的底物特异性(CAK)改变与TFIIH协会

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摘要

The transcription/DNA repair factor TFIIH consists of nine subunits, several exhibiting known functions: helicase/ATPase, kinase activity and DNA binding. Three subunits of TFIIH, cdk7, cyclin H and MAT1, form a ternary complex, cdk-activating kinase (CAK), found either on its own or as part of TFIIH. In the present work, we demonstrate that purified human CAK complex (free CAK) and recombinant CAK (rCAK) produced in insert cells exhibit a strong preference for the cyclin-dependent kinase 2 (cdk2) over a ctd oligopeptide substrate (which mimics the carboxyterminal domain of the RNA polymerase II). In contrast, TFIIH preferentially phosphorylates the ctd as well as TFIIE alpha, but not cdk2, TFIIH was resolved into four subcomplexes: the kinase complex composed of cdk7, cyclin H and MAT1; the core TFIIH which contains XPB, p62, p52, p44 and p34; and two other subcomplexes in which XPD is found associated with either the kinase complex or with the core TFIIH. Using these fractions, we demonstrate that TFIIH lacking the CAK subcomplex completely recovers its transcriptional activity in the presence of free CAK. Furthermore, studies examining the interactions between TFIIH subunits provide evidence that CAK is integrated within TFIIH via XPB and XPD. [References: 51]
机译:转录/ TFIIH包括DNA修复因素九子单元,几个展示功能:解旋酶/ atp酶激酶活性DNA结合。细胞周期蛋白H和MAT1,形成三元复杂,cdk-activating激酶(CAK),发现它自己或作为TFIIH的一部分。表明,净化人类CAK复杂(免费CAK)和重组CAK (rCAK)生产的插入细胞表现出强烈的偏好细胞周期蛋白依赖性激酶2 (cdk2) ctd寡肽底物(模仿carboxyterminal域的RNA聚合酶II)。相比之下,TFIIH优先磷酸化温盐深仪以及TFIIEα,但不是cdk2,TFIIH解决分为四个复形:细胞周期蛋白H和激酶cdk7组成的复杂MAT1;p52 p44,意思是;发现XPD与要么与核心TFIIH激酶复杂或。这些分数,我们证明TFIIH缺乏CAK复形完全恢复其转录活性的存在CAK自由。TFIIH亚基之间的相互作用证据表明CAK TFIIH通过内集成XPB和XPD。

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