Koyama N
Chemistry Department, College of Arts and Sciences, Chiba University, Japan.
Eur J Biochem. 1993 Oct 1;217(1):435-9. doi: 10.1111/j.1432-1033.1993.tb18263.x.
An anaerobic alkaliphile, EP01, specifically requires NH4+ for the acceleration of amino acid and glucose transport [Koyama, N. (1988) FEBS Lett. 253, 187-189]. In this paper, we attempted to clarify how NH4+ is involved in the transport system. The bacterium acidifies the cytoplasm, which was suggested to result in NH4+ accumulation when NH4Cl was added to the medium. Increase of the NH4Cl concentration administered to the medium caused the acceleration of leucine and glucose transport, which was accompanied by an increase in the internal pH and the absolute internal concentration of NH4+, whereas a decrease in the concentration ratio of internal NH4+/external NH4+ was observed. The addition of 3 mM NH4Cl, which resulted in significant stimulation of leucine and glucose transport, raised the internal NH4+ concentration by 42 mM, but the internal pH only by 0.1 units. It seems more likely that leucine and glucose transport are accelerated depending on the increase in the internal NH4+ concentration rather than the increase in the internal pH. By the imposition of an inwardly directed Na+ gradient, the K(+)-loaded membrane vesicles accumulated leucine and glucose, indicating that a sodium chemical potential is available for active transport. The membrane of the bacterium exhibited a Na(+)-stimulated ATPase activity which was remarkably enhanced by the addition of NH4Cl, depending on its concentration, and was inhibited by vanadate. Leucine and glucose transport were inhibited by vanadate. Based on these results, we propose a mechanism in which NH4+ contributes internally to leucine and glucose transport, depending on its concentration, by the activation of a Na(+)-translocating ATPase responsible for the generation of a sodium chemical potential.
一种厌氧嗜碱菌EP01特别需要NH₄⁺来加速氨基酸和葡萄糖的转运[小山，N.（1988年）《欧洲生物化学学会联合会快报》253，187 - 189]。在本文中，我们试图阐明NH₄⁺是如何参与转运系统的。该细菌会使细胞质酸化，当向培养基中添加NH₄Cl时，这被认为会导致NH₄⁺积累。向培养基中添加的NH₄Cl浓度增加会导致亮氨酸和葡萄糖转运加速，同时伴随着内部pH值升高以及内部NH₄⁺绝对浓度增加，而内部NH₄⁺/外部NH₄⁺的浓度比则降低。添加3 mM NH₄Cl可显著刺激亮氨酸和葡萄糖转运，使内部NH₄⁺浓度升高42 mM，但内部pH值仅升高0.1个单位。亮氨酸和葡萄糖转运似乎更有可能是依赖于内部NH₄⁺浓度的增加而加速，而非内部pH值的升高。通过施加内向的Na⁺梯度，加载了K⁺的膜囊泡积累了亮氨酸和葡萄糖，这表明钠化学势可用于主动转运。该细菌的膜表现出Na⁺刺激的ATP酶活性，添加NH₄Cl会根据其浓度显著增强该活性，且该活性受到钒酸盐的抑制。亮氨酸和葡萄糖转运受到钒酸盐的抑制。基于这些结果，我们提出一种机制，即NH₄⁺根据其浓度通过激活负责产生钠化学势的Na⁺转运ATP酶在内部促进亮氨酸和葡萄糖的转运。
