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題名:Purification and characterization of glutamine synthetase from the unicellular cyanobacterium Synechococcus RF-1     (22點)
原核聚球藻RF-1中麩醯胺合成?之純化與特性之檢測
著者:Hso-Freng Yuan(袁守方) ;Chiung-Min Wang(王瓊敏) ;Han-Wei Kung(官翰煒)
出版地區:台灣
出版城市:台北市
學科:生命科學
關鍵字:藍綠藻 ; 麩醯胺合成? ; 聚球藻RF-1 ; Cyanobacteria ; Glutamine synthetase ; Synechococcus RF-1
刊名:Botanical Bulletin of Academia Sinica
頁碼:23-33
語言:英語
摘要: 中文摘要PDF ; 英文摘要PDF

聚球藻RF-1中之麩醯胺合成?(GS; EC 6.3.1.2),可藉離子交換、分子篩濾及氫氧磷灰石(hydroxyapatite)吸著等層析步驟,純化至均一的程度。自然態的GS,其分子量為456 kDa,其次單元之分子量為56 kDa。由電子顯微鏡圖片中可以看出,該?有兩層蛋白質平行排列,以四單元一級呈立體對稱的結構。因此依實際數據,認為該?是由八個大小相同之次單元所組成。於生合成?反應中,該?對於L-Glutamate之視Km值為2.33 mM,但對於ATP(nH=1.5,S0.5=0.94 mM)及NH4Cl(nH=2,S0.5=1.33 mM)則呈現正協同性之作用。於轉移?反應中,該?對於L-Glutamine及Hydroxyamine之視Km值分別為8.70 mM及7.04 mM。該?之活性於含有EDTA、MgCl2及2-Mercaptoethanol之Tris-HCl緩衝液(pH 7.5)中非常穩定。於生合成?反應及轉移?反應中,最適於該?活性之pH值分別為8.1及8.4。該?需要兩價金屬離子來作為活性化劑;對於生合成?之活性而言,Mg2+為最有效的金屬離子,其次為Co2+;對於轉移?之活性而言,Mn2+為最有效的金屬離子。Ca2+及Mn2+對於由Mg2+離子于支持的生合成?之活性,有強烈的抑制作用,而Co2+則有促進生合成?活性之作用。
Glutamine synthetase (GS; EC 6.3.1.2) from the Synechococcus RF-1 was purified to homogeneity by ion exchange, molecular sieving, and hydroxyapatite chromatographies. The native enzyme has a molecular mass of about 456 kDa, and the molecular mass of its subunit was about 56 kDa. Electron micrographs of the enzyme revealed two parallel protein layers in cubic symmetry with quaternary structure. The actual data indicated that the enzyme could consist of eight identical subunits. The enzyme had an apparent Km value for L-glutamate of 2.33 mM, but it exhibited positive cooperativity for ATP (nH=1.5 and S0.5=0.94 mM) and NH4C1(nH=2, and S0.5=1.33 mM) in the biosynthetic assay. The enzyme had apparent Km values for L-glutamine and hydroxylamine of 8.70 mM and 7.04 mM, respectively, in the transferase assay. This enzyme was quite stable in Tris-HCI buffer (pH 7.5) containing EDTA, MgCl2, and 2-mercaptoethanol. The pH optima for both the biosynthetic and transferase activities of the enzyme were 8.1 and 8.4, respectively. The enzyme required a divalent metal ion as an activator. Mg2+ was the most effective metal ion for biosynthetic activity, followed by Co2+. Mn2+ was the most effective metal ion for transferase activity. Ca2+ and Mn2+ strongly inhibited Mg2+-supported biosynthetic activity, but Co2+ stimulated it.


    

本卷期目次
Botanical Bulletin of Academia Sinica
Adventitious shoot regeneration from stem internode explants of Adenophora triphylla (Thunb.) A. DC. (Campanulaceae)-an important medicinal herb/ Chung-Chuan ChenSuh-Jau ChenHsin-Sheng Tsay
Characteristics of the acidic environment of the Yuanyang Lake (Taiwan)/ Jiunn-Tzong WuShih-Chieh ChangYun-Sen WangYu-Fa WangMing-Kuang Hsu
Purification and characterization of glutamine synthetase from the unicellular cyanobacterium Synechococcus RF-1/ Hso-Freng YuanChiung-Min WangHan-Wei Kung
Vase life Eustoma grandiflorum as affected by aluminum sulfate/ Li-Jen LiaoYu-Han LinKuang-Liang HuangWen-Shaw Chen
Detection of leaf-associated fungi based on PCR and nucleotide sequence of the ribosomal internal transcribed spacer (ITS) in Miscanthus/ Yu-Chung ChiangChang-Hung ChouPei-Ray LeeTzen-Yuh Chiang
Mechanism of host cell death induced by infection of Escherichia coli with the c2 clear-plaque mutant of phage f1/ Shan-Hua LinWen-Ping ChenTsong-Tsong Kuo
Cold-acclimation and root temperature protection from chilling injury in chilling-sensitive mungbean (Vigna radiata L.) seedlings/ Mon-Yaw ChangSwu-Ling ChenChung-Fen LeeYih-Ming Chen
Karyomorphology of Biebersteinia Stephan (Geraniaceae) and its systematic and taxonomic significance/ Jian-Quan LiuTing-Nong HoShi-Long ChenAn-Min Lu
The genus Tubocapsicum (Solanaceae)/ Zhi-Yun ZhangChing-I Peng
High frequency somatic embryogenesis and plant regeneration of an elite Chinese cotton variety/ Bao-Hong ZhangRong FengFang LiuQing-Lian Wang
 
   
 
   

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