噬菌體 xP10 所誘導，負責關閉寄主細胞轉錄的抑制劑

Abstract

中文摘要 制劑生的由XP 10 感染的Xanthomonas oryzae細胞中，可以純化一種抑制劑，抑制寄主轉錄系統，這種抑制劑是XP10 感染寄主後，誘導產產生的。在感染 10 分鐘左右，產生量可達最高。因為與過去報告的噬菌體系統差異，為更進一步研究其特性及機制，所以將抑制劑加以純化。 感染菌以超音波細胞粉碎器打破，經硫酸銨沉澱， 60 ℃ 處理一個小時，及 Sephacryl 200 、DEAE-cellulose column chromatography 純化，可得比活性 1435 units / mg 的抑制劑，其分子量經Sephacryl 200 column chromatography測定，約在 7000－10000 delton。 抑制劑對熱相當穩定，若以60 ℃處理一個小時，可以顯著提高其純度比活性，用 protease 及 chymotrypsin 處理會完全破壞、抑制劑的活性。抑制劑明顯的只對寄主 RNA 聚合?有作用，而不影響噬菌體 RNA 聚合?，抑制劑不會直接與 DNA 作用，但可能會和寄主 RNA聚合?或〔 寄主 RNA 聚合?一 D NA 〕 複合體直接作 用。

Summary An inhibitory protein responsible for shutoff of the transcription of host cell of Xanthomonas oryzae was isolated from phage XP1O infected cells. The inhibitor is a protein and induced by phage infection. Since its properties are quite unique, therefore it was further purified and its inhibitory mechanism studied. For purification of inhibitor, the infected cells were disrupted by sonicator, the extracted inhibitor was precipitated with ammonium sulfate, the precipitate dissolved in buffer was heated at 60°C for an hour, then put through Sephacryl 200 and DEAE-cellulose column chromatography. The specific activity reached 91 folds. The inhibitor was analyzed by Sephacryl 200 column chromatography to estimate molecular weight; the obtained value was 7000-10000 daltons. The inhibitor was rather heat-stable, its specific activity increased marked during the incubation at 60°C for an hour relative to the fifteen-minutes-treated one. The activity of inhibitor is completely destroyed by incubation with protease and chymotrypsin. The inhibitor has marked effect on Xanthomonas oryzae RNA polymerase but not to the activity of XP1O specific RNA polymerase even when added at a high concentration. The inhibitor does not bind directly to DNA, but may have a direct interaction with Xanthomonas oryzae RNA polymerase or Xanthomonas oryzae RNA polymerase—DNA complex.