An N-terminal mutation in the bacteriophage T4 motA gene yields a protein that binds DNA but is defective for activation of transcription
An N-terminal mutation in the bacteriophage T4 motA gene yields a protein that binds DNA but is defective for activation of transcription
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DOI：
10.1128/jb.178.21.6133-6139.1996
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发表时间：
1996-11-01
影响因子：
3.2
通讯作者：
Hinton, DM
中科院分区：
文献类型：
--
作者：
Gerber, JS;Hinton, DM
噬菌体T4 MotA蛋白是T4修饰的宿主RNA聚合酶的转录激活因子，并且是激活中间类T4启动子所必需的，MotA单独结合T4中间启动子的-30区域，该区域含有MotA盒共有序列[（t/a）（t/a）TGCTT（t/c）A]。我们报道了命名为Mot 21的蛋白的分离和表征，其中野生型motA序列的前8个密码子已被11个不同的密码子取代。在凝胶阻滞测定中，Mot 21和MotA类似地结合含有T4中间启动子P-uvsX的DNA，并且蛋白质在P-uvsX上产生类似的足迹。然而，金属在转录激活中严重缺陷，在天然蛋白质凝胶上，在RNA聚合酶的sigma（70）亚基和野生型MotA蛋白孵育后观察到新的蛋白质种类，表明MotA和sigma（70）之间的直接蛋白质-蛋白质接触，Mot 21未能形成这种复合物，提示这种相互作用对于转录激活是必需的，并且Mot 21缺陷的出现是因为Mot 21不能像野生型激活剂那样形成这种接触。
The bacteriophage T4 MotA protein is a transcriptional activator of T4-modified host RNA polymerase and is required for activation of the middle class of T4 promoters, MotA alone binds to the -30 region of T4 middle promoters, a region that contains the MotA box consensus sequence [(t/a)(t/a)TGCTT(t/c)A], We report the isolation and characterization of a protein designated Mot21, in which the first 8 codons of the wild-type motA sequence have been replaced with 11 different codons, In gel retardation assays, Mot21 and MotA bind DNA containing the T4 middle promoter P-uvsX similarly, and the proteins yield similar footprints on P-uvsX. However, Metal is severely defective in the activation of transcription, On native protein gels, a new protein species is seen after incubation of the sigma(70) subunit of RNA polymerase and wild-type MotA protein, suggesting a direct protein-protein contact between MotA and sigma(70), Mot21 fails to form this complex, suggesting that this interaction is necessary for transcriptional activation and that the Mot21 defect arises because Mot21 cannot form this contact like the wild-type activator.