Sullivan S M, Horn P J, Olson V A, Koop A H, Niu W, Ebright R H, Triezenberg S J
Department of Biochemistry, Michigan State University, East Lansing, MI 48824-1319, USA.
Nucleic Acids Res. 1998 Oct 1;26(19):4487-96. doi: 10.1093/nar/26.19.4487.
The VP16 protein of herpes simplex virus is a potent transcriptional activator of the viral immediate early genes. The transcriptional activation region of VP16 can be divided into two functional subregions, here designated VP16N (comprising amino acids 413-456) and VP16C (amino acids 450-490). Assays of VP16C mutants resulting from both random and alanine-scanning mutagenesis indicated that the sidechains of three phenylalanines (at positions 473, 475 and 479) and one acidic residue (glutamate 476) are important for transcriptional activation. Aromatic and bulky hydrophobic amino acids were effective substitutes for each of the three Phe residues, whereas replacement with smaller or polar amino acids resulted in loss of transcriptional function. In contrast, many changes were tolerated for Glu476, including bulky hydrophobic and basic amino acids, indicating that the negative charge at this position contributes little to the function of this subregion. Similar relative activities for most of the mutants were observed in yeast and in mammalian cells, indicating that the structural requirements for this activation region are comparable in these two species. These results reinforce the hypothesis that bulky hydrophobic residues, not acidic residues, are most critical for the activity of this 'acidic' transcriptional activation region.
单纯疱疹病毒的VP16蛋白是病毒即刻早期基因的一种强效转录激活因子。VP16的转录激活区域可分为两个功能亚区，这里分别命名为VP16N（包含第413 - 456位氨基酸）和VP16C（第450 - 490位氨基酸）。对随机诱变和丙氨酸扫描诱变产生的VP16C突变体的检测表明，三个苯丙氨酸（第473、475和479位）的侧链以及一个酸性残基（谷氨酸476）对转录激活很重要。芳香族和大体积疏水氨基酸可有效替代这三个苯丙氨酸残基中的每一个，而用较小或极性氨基酸替代则导致转录功能丧失。相比之下，对于谷氨酸476可以耐受许多变化，包括大体积疏水和碱性氨基酸，这表明该位置的负电荷对该亚区的功能贡献不大。在酵母和哺乳动物细胞中观察到大多数突变体具有相似的相对活性，表明这两个物种中该激活区域的结构要求具有可比性。这些结果强化了这样一种假说，即大体积疏水残基而非酸性残基对这个“酸性”转录激活区域的活性最为关键。
