S CEREVISIAE GENERAL TRANSCRIPTION FACTOR TFIID

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 转录因子TFIID是一个多亚基复合体，在真核生物中负责将RNA聚合酶II和其他相关因子组装到启动子起始点并将其靶向和核化。它对基因表达的调控具有重要意义。基因表达的错误调节可能导致发育缺陷、代谢紊乱和癌症。了解转录启动的机制是一个重要的目标，有关TFIID的结构信息将有助于实现这一目标。TFIID复合体估计含有1.1-1.2丙二醛的蛋白质，来自15个单独的亚基。由于分离的亚基的表达和溶解性问题以及难以获得足够的材料，这一大型复合体的结构信息一直难以获得。我们的目标是扩展由P.Schultz小组使用串联亲和纯化(TAP-TAG)分离材料对酵母TFIID复合体进行的负染色电子显微镜研究。我们将获得TFIID复合体的冷冻EM重建。这些数据将用于我们获得高分辨率X射线晶体结构的努力。特别有兴趣的是使用冷冻-EM进一步表征TFIID与其他已知基本因子的相互作用，以进一步揭示RNA聚合酶II转录系统各组成部分在启动子上的空间关系。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Transcription factor TFIID is a multi-subunit complex responsible for targeting and nucleating the assembly of RNA polymerase II and other associated factors to promoter start sites in eukaryotic organisms. It is important for the regulation of gene expression. Mis-regulation of gene expression can cause developmental defects, metabolic disorders, and cancer. Understanding the mechanism of transcriptional initiation is an important goal, and structural information about TFIID would contribute towards this end. The TFIID complex is estimated to contain 1.1-1.2 MDa of protein contributed from 15 individual subunits. Structural information for this large complex has been difficult to obtain due to expression and solubility problems for the isolated subunits and difficulty in obtaining sufficient material. Our goal has been to extend on negative stain electron microscopy studies of the yeast TFIID complex carried out by the group of P. Schultz using Tandem Affinity Purification (TAP-tag) isolated material. We will obtain cryo-EM reconstructions of the TFIID complex. This data will be used in our efforts to obtaining a high-resolution X-ray crystal structure. It would be of particular interest to further characterize the interactions of TFIID with other known basal factors using cryo-EM to further reveal the spatial relationships between the various components of the RNA Polymerase II transcription system at promoters.