完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.author | 游金珠 | en_US |
dc.contributor.author | Yu, Chin-Chu | en_US |
dc.contributor.author | 曾 慶 平, 黃 效 民, 1, 1, 1, 1 | en_US |
dc.contributor.author | Tseng Ching-Ping, Hwang Shiaw-Min, 1, 1, 1, 1 | en_US |
dc.date.accessioned | 2014-12-12T02:14:38Z | - |
dc.date.available | 2014-12-12T02:14:38Z | - |
dc.date.issued | 1995 | en_US |
dc.identifier.uri | http://140.113.39.130/cdrfb3/record/nctu/#NT840111007 | en_US |
dc.identifier.uri | http://hdl.handle.net/11536/60075 | - |
dc.description.abstract | 中文摘要 延胡索酸酵素 (fumarase) 為三梭 梭酸循環 (tricaboxylic acid cycle, TCA cycle) 中重要之酵素, 其作 用為催化延胡索酸 (fumarate)與蘋果酸 (malate) 之間的轉換. 大腸桿 菌 (Escherichia coli) 含有三種能產生延胡索酸酵素的基因, 分別命名 為 fumA, fumB 及 fumC. 就酵素的生化特異性而言, fumA 與 fumB 基因 所產生的 fumarase ( FumA, fumB) 為熱不安定性 (heat labile) 酵素, 其蛋白質含有鐵硫活性中心. 而 fumC 的基因產物 (FumC) 則為熱安定 (heat stable) 酵素, 其酵素活性反應無需鐵離子參與, 而且 fumC 基因 的表現會受 superoxide (O2-) 誘發. 為了研究細胞內 fumA 基因的 表現與生長速率的關係, 我們利用分批式培養法及化學恆定 (chemstat) 連續式培養法來控制細胞的生長速率, 以探討不同生長速率下 fumA 基因 的表現量. 由實驗結果發現, 生長速率與 FumA 活性的關係在批式培養 時, 當生長速率變慢時, FumA 的活性上升. 而在連續式培養中, 我們發 現改變細胞的生長速率, 所測得 FumA 的活性並無顯著變化. 另外, 在連 續式培養時 FumC 的活性會隨生長速率得下降而增加. 當我們控制細 胞的生長速率由慢變快時, 分別抽取細胞在不同生長速率下的總 RNA,然 後進行北方墨點轉印分析, 結果顯示在分批式與連續式培養中, fumA 基 因所產生的mRNA 隨生長速率的上升而產量均下降. 此結果表示 fumA 基 因的表現, 在 DNA 轉錄為mRNA (transcriptional level) 的步驟已受生 長速率的影響. 由上述實驗結果使我們認為有必要繼續探討 fumA 基因在 不同生長速率下所產生的 mRNA 之穩定性. 利用 rifampin 抑制 DNA 轉 錄為 mRNA 的啟始步驟, 進而測定不同生長速率下 fumA mRNA 之半衰期. 結果顯示 fumA 基因產生的 mRNA 在比生長速率為 0.48/hr 時, 半衰期 為 0.67 分鐘, 在比生長速率為 1.2/hr 時, 半衰期延長為 3.99 分鐘. 所以當細胞生長速率愈快, 會使 fumA 基因產生的 mRNA 愈穩定. 綜合上 述結果我們可知, 控制大腸桿菌細胞的生長速率可調控 fumA 基因的表 現, 此調控機制主要發生在 transcriptional (mRNA 的量) 及 post- transcriptional level (mRNA 的穩定性). Abstract The tricarboxylic acid cycle enzyme fumarasecatalyzes the interconversion of fumarate and L-malate. Three biochemically distinctfumarases have been reported in Escherichia coli. While the fumA and fumBgenes encode heat-labile, iron containing fumarase, the fumC gene product is a heat-stable fumarase which does not require iron for activity. The expressionof fumC gene is regulated by superoxide (O2-) levels though a regulon called soxRS. To study the relation between the expression of fumA gene and cell growth rate in E. coli, we used batch and continuous culture to control cellgrowth rate. The results showed that the activity of FumA decreased when cellgrowth rate was increasd in batch culture, however, the activity of FumA did not apparently change with cell growthrate in continuous culture. The activity of FumC decreased 4-fold as cellgrowth rate increased 5-fold. With Northern (RNA) analysis, we found that in both batch culture and continuous culture the amount of fumA mRNA relative to total RNA decreased wiyh increasing cell growth rate. This result indicated that the growth rate-dependent regulation happened at the transcriptional level. In addition, we measured the decay of fumA mRNA after blocking transcription initiation with rifampin and found that the stability of fumA mRNA increased about 6-fold as cell growth rate increased 2.5-fold. These results suggested that growth rate-dependent regulation of fumA gene was mediated by both transcriptional and post-transcriptional mechanism. | zh_TW |
dc.language.iso | zh_TW | en_US |
dc.subject | 延胡索酸酵素 | zh_TW |
dc.subject | 大腸桿菌 | zh_TW |
dc.subject | 生長速率 | zh_TW |
dc.subject | 連續式培養 | zh_TW |
dc.subject | 轉錄 | zh_TW |
dc.subject | 北方墨點轉印 | zh_TW |
dc.subject | fumarase | en_US |
dc.subject | Escherichia coli | en_US |
dc.subject | growth rate | en_US |
dc.subject | continuous culture | en_US |
dc.subject | transcription | en_US |
dc.subject | northern blotting | en_US |
dc.title | 細胞生長速率對大腸桿菌延胡索酸酵素基因表現之影響 | zh_TW |
dc.title | Effect of cell growth rate on fumarase gene fumA in E. coli | en_US |
dc.type | Thesis | en_US |
dc.contributor.department | 生物科技學系 | zh_TW |
顯示於類別: | 畢業論文 |