Speelmans G, de Vrij W, Konings W N
Department of Microbiology, University of Groningen, Haren, The Netherlands.
J Bacteriol. 1989 Jul;171(7):3788-95. doi: 10.1128/jb.171.7.3788-3795.1989.
Amino acid transport was studied in membrane vesicles of the thermophilic anaerobic bacterium Clostridium fervidus. Neutral, acidic, and basic as well as aromatic amino acids were transported at 40 degrees C upon the imposition of an artificial membrane potential (delta psi) and a chemical gradient of sodium ions (delta microNa+). The presence of sodium ions was essential for the uptake of amino acids, and imposition of a chemical gradient of sodium ions alone was sufficient to drive amino acid uptake, indicating that amino acids are symported with sodium ions instead of with protons. Lithium ions, but no other cations tested, could replace sodium ions in serine transport. The transient character of artificial membrane potentials, especially at higher temperatures, severely limits their applicability for more detailed studies of a specific transport system. To obtain a constant proton motive force, the thermostable and thermoactive primary proton pump cytochrome c oxidase from Bacillus stearothermophilus was incorporated into membrane vesicles of C. fervidus. Serine transport could be driven by a membrane potential generated by the proton pump. Interconversion of the pH gradient into a sodium gradient by the ionophore monensin stimulated serine uptake. The serine carrier had a high affinity for serine (Kt = 10 microM) and a low affinity for sodium ions (apparent Kt = 2.5 mM). The mechanistic Na+-serine stoichiometry was determined to be 1:1 from the steady-state levels of the proton motive force, sodium gradient, and serine uptake. A 1:1 stoichiometry was also found for Na+-glutamate transport, and uptake of glutamate appeared to be an electroneutral process.
对嗜热厌氧细菌嗜热栖热梭菌的膜囊泡中的氨基酸转运进行了研究。在施加人工膜电位（Δψ）和钠离子化学梯度（ΔμNa⁺）的情况下，中性、酸性、碱性以及芳香族氨基酸在40℃时被转运。钠离子的存在对于氨基酸的摄取至关重要，仅施加钠离子化学梯度就足以驱动氨基酸摄取，这表明氨基酸是与钠离子协同转运而非与质子协同转运。锂离子，但不是所测试的其他阳离子，可在丝氨酸转运中替代钠离子。人工膜电位的瞬态特性，尤其是在较高温度下，严重限制了它们在更详细研究特定转运系统中的适用性。为了获得恒定的质子动力，将来自嗜热脂肪芽孢杆菌的热稳定且热活性的初级质子泵细胞色素c氧化酶整合到嗜热栖热梭菌的膜囊泡中。丝氨酸转运可由质子泵产生的膜电位驱动。离子载体莫能菌素将pH梯度转化为钠离子梯度刺激了丝氨酸摄取。丝氨酸载体对丝氨酸具有高亲和力（Kt = 10 μM），对钠离子具有低亲和力（表观Kt = 2.5 mM）。根据质子动力、钠离子梯度和丝氨酸摄取的稳态水平，确定Na⁺ - 丝氨酸的机制化学计量比为1:1。对于Na⁺ - 谷氨酸转运也发现了1:1的化学计量比，并且谷氨酸的摄取似乎是一个电中性过程。
