Kadonaga J T, Carner K R, Masiarz F R, Tjian R
Howard Hughes Medical Institute, Department of Biochemistry, University of California, Berkeley 94720.
Cell. 1987 Dec 24;51(6):1079-90. doi: 10.1016/0092-8674(87)90594-0.
Transcription factor Sp1 is a protein present in mammalian cells that binds to GC box promoter elements and selectively activates mRNA synthesis from genes that contain functional recognition sites. We have isolated a cDNA that encodes the 696 C-terminal amino acid residues of human Sp1. By expression of truncated fragments of Sp1 in E. coli, we have localized the DNA binding activity to the C-terminal 168 amino acid residues. In this region, Sp1 has three contiguous Zn(II) finger motifs, which are believed to be metalloprotein structures that interact with DNA. We have found that purified Sp1 requires Zn(II) for sequence-specific binding to DNA. Thus, it is likely that Sp1 interacts with DNA by binding of the Zn(II) fingers. To facilitate the identification of mutant variants of Sp1 that are defective in DNA binding, we have also devised a bacterial colony assay for detection of Sp1 binding to DNA.
转录因子Sp1是一种存在于哺乳动物细胞中的蛋白质，它与GC盒启动子元件结合，并选择性地激活含有功能性识别位点的基因的mRNA合成。我们分离出了一个编码人Sp1 C末端696个氨基酸残基的cDNA。通过在大肠杆菌中表达Sp1的截短片段，我们将DNA结合活性定位到C末端的168个氨基酸残基。在这个区域，Sp1有三个相邻的锌（II）指基序，据信它们是与DNA相互作用的金属蛋白结构。我们发现纯化的Sp1需要锌（II）才能与DNA进行序列特异性结合。因此，Sp1很可能通过锌（II）指的结合与DNA相互作用。为了便于鉴定在DNA结合方面有缺陷的Sp1突变变体，我们还设计了一种细菌菌落检测法来检测Sp1与DNA的结合。
