聚球藻RF-1硝酸鹽代謝的特性研究

Abstract

聚球藻Synechococcus RF-1?具固氮能力且被發現有概日韻律之單細胞藍綠藻，但在硝酸鹽?氮源時，其生長更?迅速。本論文以其?材料，探討其利用硝酸鹽之特性，經分析硝酸鹽之吸收，及硝酸還原酵素活性變化的特性，以探討其硝酸還原反應的調控情形。結果發現，在無硝酸鹽之培養下，細胞仍具硝酸還原酵素之活性，顯示，硝酸還原酵素之活性不需要硝酸鹽之誘導。在光／暗週期生長下，硝酸還原酵素活性未發現有高低起伏之韻律現象。在含有硝酸鹽的培養條件下，添加氯黴素時，酵素活性衰減之速率也很小，此顯示，聚球藻RF-1之硝酸還原酵素非常穩定。聚球藻對硝酸鹽的吸收在光／暗週期下吸收量變化很大，當自光／暗週期培養下移至全光照下，吸收量仍會隨原來之韻律期而起伏。顯然硝酸鹽之吸收步驟，才是調控硝酸鹽利用的部位，而非硝酸還原酵素。當以穀胺醯胺合成之抑制劑MSX處理培養於不同氮源之藻細胞，結果顯示，銨離子不會抑制NR活性，但會抑制硝酸根離子之利用，經進一步添加Gln及MSX試驗証明，銨離子非直接抑制硝酸還原反應之進行，而是經由其合成物穀胺醯胺而達成抑制作用，且需銨與穀胺醯胺共同作用，才會影響硝酸鹽之吸收與硝酸鹽之還原反應。

Synechococcus RF-1 is an N2-fixing unicellular cyanobacterium which is found to have circadian rhythm of N2-fixing activity. Present study is aimed to characterize the utilization of nitrate and the control step of this process. The kinetics of the uptake of nitrate and of the enzyme nitrate reductase were studied. It was revealed that both of the cells grown in the presence or absence of nitrate did have approximately the same NR activity, indicating that nitrate is not an obligate inducer for NR. In the cells grown under L/D cycle, the NR activity remained nearly unchanged during a cycle. The turnover rate of NR was very low even when chloram-phenicol was added to the cultures cultivated with nitrate as N-source. This implies that the NR in Synechococcus RF-1 is rather stable in the cells. The comparison of the uptake of nitrate in the cells grown in the light with that grown in the dark shows that there is an ossillation in the magnitude of nitrate uptake during a L/D cycle. The cells keep the rhythmic ossillation of nitrate uptake even when they were transfered to a LL conditions. All of the results indicate that it is the uptake of nitrate rather than the assimilation process which regulates the ossillation of nitrate utilization. Ammonium is an effective inhibitor for nitrate utilization. It was found that ammonium did not inhibit NR activity directly. Rather it inhibited the uptake of nitrate via its product, glutamine, like some of other Cyanophyte.