Westin G, Schaffner W
Institut für Molekularbiologie II, Universität Zürich, Switzerland.
Nucleic Acids Res. 1988 Jul 11;16(13):5771-81. doi: 10.1093/nar/16.13.5771.
Zinc is an important cofactor for many enzymes involved in nucleic acid metabolism such as DNA and RNA polymerases, reverse transcriptase and tRNA synthetases. We have developed an inducible in vitro transcription system using metal-depleted nuclear extracts to reveal the presence and functional relevance of heavy metal ions in transcription factors. Using protein-DNA binding assays (band shift and DNAase I footprint) we show that Sp1, a promoter-specific vertebrate transcription factor that binds to the "GC box" (Sequence in text), is reversibly inactivated by metal-depletion. Zinc is required for specific DNA binding in vitro and is also essential for Sp1 factor-directed transcription. In contrast, another factor from HeLa cells, the so-called octamer transcription factor (OTF) that binds to the sequence 5'-ATGCAAATNA, is not affected by metal-depletion and thus seems not to be a zinc metalloprotein.
锌是许多参与核酸代谢的酶的重要辅助因子，如DNA和RNA聚合酶、逆转录酶和tRNA合成酶。我们利用金属耗尽的核提取物开发了一种可诱导的体外转录系统，以揭示重金属离子在转录因子中的存在及其功能相关性。通过蛋白质-DNA结合分析（凝胶迁移和DNA酶I足迹法），我们发现Sp1，一种与“GC盒”（文中序列）结合的启动子特异性脊椎动物转录因子，会因金属耗尽而可逆失活。锌是体外特异性DNA结合所必需的，也是Sp1因子指导转录所必需的。相比之下，来自HeLa细胞的另一种因子，即与序列5'-ATGCAAATNA结合的所谓八聚体转录因子（OTF），不受金属耗尽的影响，因此似乎不是一种锌金属蛋白。
