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標題: 嗜鹽性甲烷菌Methanohalophilus portucalensis運送glycine betaine 的機制
Glycine betaine transport in the Methanohalophilus portucalensis
作者: 洪堂耀
Hong, Tang-Yao
關鍵字: glycine betaine;嗜鹽性甲烷菌;transport;kinetic;運送;動力學
出版社: 植物學系
Methanohalophilus portucalensis FDF1是一株中度嗜鹽性甲烷古生菌,
最適合生長的鹽濃度範圍在1.2-2.9 M之間,其對高鹽環境的適應機制,
Ectothiorhodospira halochloris、Aphanothece halophytica及嗜鹽性
甲烷菌是已知可自體生合成glycine betaine的微生物,其它微生物大多
自環境中攝取glycine betaine或其前驅物choline到細胞內累積。有關
glycine betaine攝取機制的研究,在真細菌中已有廣泛報導,但嗜鹽性
甲烷菌攝取glycine betaine的機制則不清礎。本實驗利用choline
oxidase氧化[14C]choline的方法製備[14C]glycine betaine,探討有關
Mh. portucalensis FDF1攝取glycine betaine的機制。在含2.1 M NaCl
沒有添加碳源的液體培養基中,此菌株對glycine betaine運輸之Km
為23.1 *M, Vmax為8.0 nmole‧min-1‧mg protein-1,屬於高親和性的
運輸系統。當受高鹽刺激時,於1.5-2.5 M NaCl間其攝取速率隨鹽濃度的
增加而提高,顯示glycine betaine的攝取受到滲透壓的調控。利用代謝
株的ATP生成,結果對glycine betaine的攝取有明顯的抑制,證明了此菌
株glycine betaine運輸系統為ATP-dependent。在專一性測試方面,
proline、choline及N,N-dimethylglycine對glycine betaine的攝取速率
並沒有影響,而sarcosine、glycine各有26% 及29%抑制,glycine
betaine有高達94%的抑制作用,顯示此glycine betaine運輸系統具有很
高的專一性。由上述結果可知Mh. portucalensis FDF1 glycine betaine
Methanohalophilus portucalensis FDF1 can grow optimally over the
salt range of 1.2-2.9 M. In response to the high salt
environment, Mh. portucalensis FDF1 synthesize glycine betaine,
b-glutamine, and Ne-acetyl-b-lysine as compatible solutes. With
the addition of glycine betaine in the medium, the doubling time
of this glycine betaine synthesizing archaea decreasedand the
biosynthesis of compatible solutes were repressed which both
indicated the occurrence of glycine betaine transport system in
Mh. portucalensis FDF1. Assay of the transport of glycine
betaine were performed with the addition of [14C]glycine betaine
which was preparedby the oxidation of [14C]choline. The
transport kinetic of Mh. portucalensis FDF1 exhibitedthe
Michaelis-Menten kinetics, with Km value of 23.1 *M and Vmax of
8.0 nmole . min-1 . mg protein-1. The uptake rate of glycine
betaine increased while the cells encountered to salt shock
within the salt range of 1.5-2.5 M and dropped at higher salt
concentration. The uncoupler 2,4-dinitrophenol and ATPase
inhibitor N,N-dicyclohexylcarbodiimide blocked the uptake of
glycine betaine; however the cytochrome oxidase inhibitor,
sodium azide, had no effect on this transport system. Besides,
this transport system was highly specific; neitherproline,
choline, nor N,N-dimethylglycine competed effectively with
glycine betaine. These results implied that Mh. portucalensis
strain FDF1 possesses an active, specific and osmoticstress
regulated glycine betaine transport system.
Appears in Collections:生命科學系所

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