Waldschmidt R, Jahn D, Seifart K H
Institut für Molekularbiologie und Tumorforschung, Marburg/Lahnberge, Federal Republic of Germany.
J Biol Chem. 1988 Sep 15;263(26):13350-6.
Transcription factor IIIB (TFIIIB), which by itself does not bind stably or specifically to DNA, was purified from cytoplasmic extracts of HeLa cells using five different chromatographic steps. This procedure yields one predominant polypeptide which represents 90% of the most highly purified preparation and shows a relative molecular mass of 60,000, when analyzed on sodium dodecyl sulfate-polyacrylamide gels. A similar value was obtained for the native protein by rate zonal centrifugation on glycerol gradients. From these data we conclude that TFIIIB from HeLa cells has a Mr of 60,000 +/- 5,000 and that it functions as a single polypeptide. Highly purified TFIIIB was required and sufficient for the specific transcription of the Xenopus laevis and human tRNA and 5 S RNA genes as well as those for VA RNA when reconstituted with RNA polymerase III and the other appropriate transcription factors.
转录因子IIIB（TFIIIB）本身不能稳定或特异性地结合DNA，它是通过五个不同的色谱步骤从HeLa细胞的细胞质提取物中纯化出来的。该方法产生一种主要的多肽，在十二烷基硫酸钠-聚丙烯酰胺凝胶上分析时，它占最高纯度制剂的90%，相对分子质量为60,000。通过在甘油梯度上进行速率区带离心，天然蛋白也获得了类似的值。根据这些数据，我们得出结论，HeLa细胞中的TFIIIB的相对分子质量为60,000±5,000，并且它作为单一多肽发挥作用。当与RNA聚合酶III和其他合适的转录因子重构时，高度纯化的TFIIIB对于非洲爪蟾和人类tRNA及5S RNA基因以及VA RNA基因的特异性转录是必需且足够的。
