Purification and Characterization of RNA polymerase from halophilic archaebacteria.

盐生盐杆菌（Halobacterium halobium）是嗜盐古细菌之一，生长在极端嗜盐环境中。细胞内的K^+浓度也非常高。盐生盐藻基因转录的机制以及参与这一过程的RNA聚合酶的性质还有待进一步研究。我们试图纯化需要高盐浓度才能发挥作用的RNA聚合酶。Zilling等（1978）从一种相关的H.cutirubrum中鉴定出一种高分子量RNA聚合酶。在H.halobium细胞的粗提物中，我们检测到两种RNA聚合酶活性，一种需要高浓度的盐才能发挥最大活性，另一种不需要。高浓度的RNA聚合酶被纯化至约1，经PEI迭代、Buty-Toyopolymer和甘油梯度离心，部分纯化的RNA聚合酶需要3 M NaCl的最大活性DNA依赖的RNA合成至少持续到90分钟，在40 ℃。该酶是耐利福平和α-鹅膏蕈碱，但敏感的木质素，这是一种有效的抑制剂流感病毒RNA聚合酶。甘油梯度离心法测得其分子量约为45 kD。酶的进一步纯化和酶结构的详细分析正在进行中。

Halobacterium halobium, one of the halophilic archaebacteria, grows in extreme halophilic environments. The intracellular concentration of K^+ is also extremely high. The mechanisum of gene transcription in H.halobium and the nature of RNA polymerase involved in this process are still remained to study. We tried to purify the RNA polymerase which requires high salt concentrations for function.Zilling et al. (1978) identified a high-molecular-weight RNA polymerase from a related H.cutirubrum.In crude extract of H.halobium cells, we detected two RNA polymerase activities, one requiring high concentrations of salt for maximun activity and the other not requiring it. The high-requiring RNA polymerase was purified up to about 1,000-fold by PEI iteatment, Buty-Toyopearl and glycerol gradient centrifugation. The partially purified RNA polymerase required 3M NaCl for maximum activity DNA-dependent RNA synthesis continued at least until 90 min at 40ﾟC.The enzyme was resistant to both rifampicin and alpha-amanitin, but was sensitive to lignin, which is a potent inhibitor of influenza virus RNA polymerase. The molecular weight was estimated to be about 45 kDs by glycerol gradient centrifugation. Futher purification of the enzyme and detailed analysis of the enzyme structure are in progress.