Response of Bacillus subtilis to high osmolarity:: uptake of carnitine, crotonobetaine and γ-butyrobetaine via the ABC transport system OpuC
Response of Bacillus subtilis to high osmolarity:: uptake of carnitine, crotonobetaine and γ-butyrobetaine via the ABC transport system OpuC
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DOI：
10.1099/00221287-144-1-83
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发表时间：
1998-01-01
期刊：
影响因子：
--
通讯作者：
Bremer, E
中科院分区：
文献类型：
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作者：
Kappes, RM;Bremer, E
研究发现，低浓度的天然甜菜碱-肉碱、巴豆甜菜碱和γ-丁基甜菜碱对枯草芽孢杆菌具有高度的渗透耐受性。对微量培养细胞摄取L-[N-甲基-C-14]肉碱的动力学分析表明，该转运系统存在一个高亲和力转运系统，其K-m值为5µM，最大转运速率(V-max)为41nmolmin(-1)(mg蛋白)(-1)。对一组携带甘氨酸甜菜碱运输系统OpuA、OpuC和OpuD明确突变的菌株的生长和运输研究表明，三磷酸腺苷结合盒运输系统OpuC是枯草杆菌摄取L肉碱的唯一途径。与巴豆甜菜碱和γ-丁基甜菜碱的竞争实验表明，OpuC体系对这些三甲基铵化合物也表现出很高的亲和力，K-I值为6.4m。放射性标记的L肉碱的示踪实验和细胞提取物的C-13-核磁共振示踪表明，这些甜菜碱是由枯草杆菌以未经修饰的形式积累的。相反，β-取代的乙酰肉碱和辛酰肉碱对枯草杆菌具有渗透保护剂的作用，但在细胞中以肉碱的形式积累。这些三甲基铵化合物都不是唯一的碳源或氮源。这些结果表明L-肉碱、巴豆甜菜碱和γ-丁基甜菜碱是枯草杆菌的有效相容溶质，并确立了ABC转运系统OpuC在枯草杆菌供应各种渗透保护剂中的关键作用。
It was found that low concentrations of the naturally occurring and structurally related betaines L-carnitine, crotonobetaine and gamma-butyrobetaine conferred a high degree of osmotic tolerance to Bacillus subtilis. Kinetic analysis of L-[N-methyl-C-14]carnitine uptake in cells grown in minimal medium revealed the presence of a high-affinity transport system with a K-m value of 5 mu M and a maximum rate of transport (V-max) of 41 nmol min(-1) (mg protein)(-1) A rise in medium osmolarity moderately increased the maximum velocity [V-max 71 nmol min(-1) (mg protein)(-1)] of this transport system, but had little effect on its affinity. Growth and transport studies with a set of strains that carried defined mutations in the previously identified glycine betaine transport systems OpuA, OpuC and OpuD allowed the identification of the ATP-binding cassette (ABC) transport system OpuC as the only uptake route for L-carnitine in B. subtilis. Competition experiments with crotonobetaine and gamma-butyrobetaine revealed that the OpuC system also exhibited a high affinity for these trimethylammonium compounds with K-i values of 6.4 mu M. Tracer experiments with radiolabelled L-carnitine and C-13-NMR tracings of cell extracts demonstrated that these betaines are accumulated by B. subtilis in an unmodified form. In contrast, the beta-substituted acylcarnitine esters acetylcarnitine and octanoylcarnitine both functioned as osmoprotectants for B. subtilis but were found to be accumulated as carnitine by the cells. None of these trimethylammonium compounds were used as sole carbon or nitrogen sources. The results thus characterize L-carnitine, crotonobetaine and gamma-butyrobetaine as effective compatible solutes for B. subtilis and establish a crucial role of the ABC transport system OpuC for the supply of B. subtilis with a variety of osmoprotectants.