Schelcher Celine, Al Mehairi Salama, Verrall Elizabeth, Hope Questa, Flower Kirsty, Bromley Beth, Woolfson Derek N, West Michelle J, Sinclair Alison J
School of Life Sciences, University of Sussex, Brighton BN1 9QG, United Kingdom.
J Virol. 2007 Jul;81(13):7149-55. doi: 10.1128/JVI.00215-07. Epub 2007 Apr 25.
The Epstein-Barr virus transcription factor Zta (encoded by BZLF1) is a bZIP protein containing an alpha-helical coiled-coil homodimerization motif (zipper). The Zta zipper forms less-stable dimers than other bZIP proteins, and an adjacent region (CT) interacts with the zipper to form a novel structure that is proposed to strengthen the dimer. Here we question the role of the CT region for Zta function. Cross-linking experiments demonstrate that the entire CT region lies adjacent to the zipper. Detailed analyses of Zta truncation mutations identify an involvement of the proximal CT region (221 to 230) in dimer formation with a further contribution from the distal region (236 to 243). Biophysical analyses reveal that residues 221 to 230 enhance the stability of the coiled coil. The ability of the Zta truncation mutants to interact with three Zta-binding sites also requires the proximal CT region. Fine mapping of DNA-binding requirements highlighted the contribution of these amino acids for Zta function. Thus, the proximal part of the CT region is required to aid the dimerization of Zta and thereby its DNA-binding ability. In contrast, although the distal part of the CT region aids dimerization, it promotes only a modest increase in DNA binding. To probe this further, we defined the contribution from the CT region for Zta to transactivate a promoter embedded within the viral genome. From this we conclude that the proximal part of the CT region is absolutely required, whereas the distal part is dispensable.
爱泼斯坦-巴尔病毒转录因子Zta（由BZLF1编码）是一种bZIP蛋白，含有一个α-螺旋卷曲螺旋同二聚化基序（拉链）。与其他bZIP蛋白相比，Zta拉链形成的二聚体稳定性较低，且一个相邻区域（CT）与拉链相互作用形成一种新结构，该结构被认为可加强二聚体。在此，我们对CT区域在Zta功能中的作用提出质疑。交联实验表明，整个CT区域与拉链相邻。对Zta截短突变体的详细分析确定，近端CT区域（221至230）参与二聚体形成，远端区域（236至243）也有进一步贡献。生物物理分析表明，221至230位残基增强了卷曲螺旋的稳定性。Zta截短突变体与三个Zta结合位点相互作用的能力也需要近端CT区域。对DNA结合需求的精细定位突出了这些氨基酸对Zta功能的贡献。因此，CT区域的近端部分是Zta二聚化及其DNA结合能力所必需的。相比之下，尽管CT区域的远端部分有助于二聚化，但它仅使DNA结合适度增加。为进一步探究此问题，我们确定了CT区域对Zta激活病毒基因组内启动子的贡献。由此我们得出结论，CT区域的近端部分是绝对必需的，而远端部分则是可有可无的。
