PATTERN OF AROMATIC AND HYDROPHOBIC AMINO-ACIDS CRITICAL FOR ONE OF 2 SUBDOMAINS OF THE VP16 TRANSCRIPTIONAL ACTIVATOR
PATTERN OF AROMATIC AND HYDROPHOBIC AMINO-ACIDS CRITICAL FOR ONE OF 2 SUBDOMAINS OF THE VP16 TRANSCRIPTIONAL ACTIVATOR
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DOI：
10.1073/pnas.90.3.883
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发表时间：
1993-02-01
影响因子：
11.1
通讯作者：
TRIEZENBERG, SJ
中科院分区：
文献类型：
--
作者：
REGIER, JL;SHEN, F;TRIEZENBERG, SJ
通过寡核苷酸定向诱变检查了单纯疱疹病毒体蛋白 VP16 转录激活域的结构特征。截短的 VP16 激活结构域 (DELTA456) 的 442 位（通常由苯丙氨酸残基占据）进行了广泛的诱变，证明了该位置芳香族氨基酸的重要性。基于Phe-442周围的VP16序列和其他转录激活结构域的序列的比对，我们对VP16的位置439和444处的亮氨酸残基进行诱变。这些实验的结果表明，Phe-442 侧翼的大疏水残基也对截短的 VP16 激活结构域的功能有显着贡献。将氨基酸 457-490 恢复到各种 Phe-442 突变体部分恢复了活性。尽管 Phe-473 周围的氨基酸模式与 Phe-442 周围的氨基酸模式相似，但 Phe-473 的突变并没有显着影响活性；事实上，Phe-475 似乎比 Phe-473 对突变更敏感。我们推断 VP16 的两个区域（氨基酸 413-456 和 457-490）具有独特的结构特征，尽管它们都不太可能是两亲性 α 螺旋或“酸性斑点”。结合之前的体外激活和抑制研究，这些结果表明 VP16 的两个子结构域通过不同的机制影响转录。
Structural features of the transcriptional activation domain of the herpes simplex virion protein VP16 were examined by oligonucleotide-directed mutagenesis. Extensive mutagenesis at position 442 of the truncated VP16 activation domain (DELTA456), normally occupied by a phenylalanine residue, demonstrated the importance of an aromatic amino acid at that position. On the basis of an alignment of the VP16 sequence surrounding Phe-442 and the sequences of other transcriptional activation domains, we subjected leucine residues at positions 439 and 444 of VP16 to mutagenesis. Results from these experiments suggest that bulky hydrophobic residues flanking Phe-442 also contribute significantly to the function of the truncated VP16 activation domain. Restoration of amino acids 457-490 to various Phe-442 mutants partially restored activity. Although the pattern of amino acids surrounding Phe-473 resembles that surrounding Phe-442, mutations of Phe-473 did not dramatically affect activity; in fact, Phe-475 appears more sensitive to mutations than does Phe-473. We infer that the two regions of VP16 (amino acids 413-456 and 457-490) possess unique structural features, although neither is likely to be an amphipathic alpha-helix or an ''acidic blob.'' These results, considered with previous in vitro activation and inhibition studies, suggest that the two subdomains of VP16 affect transcription by different mechanisms.