STRUCTURAL CHARACTERIZATION OF THE TAF1-TAF7 TFIID SUBCOMPLEX

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 转录因子TFIID由TATA盒结合蛋白(TBP)和14个TBP相关因子(TAFs)组成，在RNA聚合酶II的基因表达调控中起着关键作用。TFIID作为核心启动子识别因子在真核生物蛋白质编码基因起始点的转录装置组装过程中起着关键作用。对TFIID复合体的各种生化研究表明，该复合体最大的亚基TAF1具有组蛋白乙酰转移酶和激酶活性，这可能在TFIID依赖启动子的基因表达调控中发挥关键作用。虽然这些活性已经被推测，但TAF1结构域与任何已知的激酶或乙酰基转移酶家族成员没有序列同源性。尽管TAF1的一些结构域和功能已经被鉴定，但TAF1分子中最大的C-末端区域被确定为可能的酶结构域仍然未知。因此，我们有兴趣获得TFIID的TAF1亚单位的结构信息。我们以前已经鉴定了一个稳定的TFIID亚复合体，它由TAF1亚单位的催化区域和TAF7的调节区组成，并确定了从大肠杆菌中表达和纯化该复合体的条件。我们希望通过结晶学的方法确定TFIID的TAF1-TAF7亚复合体的结构，并相信这将为我们提供第一次对预引发复合体的组织和组装的结构一瞥。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The transcription factor TFIID, composed of the TATA box binding protein (TBP) and 14 TBP associated factors (TAFs), plays a key role in regulation of gene expression by RNA Polymerase II. TFIID plays a critical role as a core promoter recognition factor that begins the process of assembling the transcriptional apparatus at the start site of protein coding genes in eukaryotes. A variety of biochemical studies of TFIID complexes have suggested that the largest subunit of the complex, TAF1, possesses histone acetyl-transferase and kinase activities that may play critical roles in the regulation of gene expression from TFIID dependent promoters. While these activities have been postulated, TAF1 domains possess no sequence homology with any known kinase or acetyl-transferase family members. Although some TAF1 domains and functions have been characterized, the largest C-terminal region of the TAF1 molecule identified as the likely enzymatic domain remains structurally unknown. Consequently we are interested in obtaining structural information for the TAF1 subunit of TFIID. We have previously identified a stable subcomplex of TFIID that consists of the putative catalytic region of the TAF1 subunit and the regulatory region of TAF7 and have determined conditions for the expression and purification of this complex from E. coli. We hope to determine the structure of the TAF1-TAF7 subcomplex of TFIID crystallographically and believe this will provide the first structural glimpse into the organization and assembly of the preinitiation complex.