MOLECULAR STRUCTURE AND PHYSIOLOGICAL CHANGES OF MEMBRANE TRANSPORT SYSTEMS OF PLANT VACUOLE.

我们研究了植物液泡的膜运输系统。液泡执行许多对细胞内稳态至关重要的功能，包括代谢物和离子的积累。这些液泡功能由初级和次级主动运输系统支持。两个质子泵，即H^+-ATP酶和H^+-焦磷酸酶，酸化液泡并产生膜电位，膜电位被用作次级运输系统的能量来源。除了这两个质子泵外，我们还分析了一个非常疏水的蛋白质，暂命名为VM 23，它被认为是液泡膜的水通道。(1)H^+-焦磷酸酶广泛存在于蕨类、苔藓等绿色植物中。因此，H^+-焦磷酸酶是绿色植物中的一种常见酶。然而，VM 23在伞藻属、轮藻属、锥头藻属和卡氏藻属的液泡膜中没有发现。(2)我们得到了大麦H^+-焦磷酸酶的cDNA。在从cDNA估计的一级氨基酸序列中，我们发现了两个重要部分，底物水解的催化部分和质子通道形成结构域。这一信息支持我们的建议，一个单一的多肽73千道尔顿不仅水解无机焦磷酸盐，但也运输质子跨膜。(3)在南瓜子叶萌发过程中，蛋白体转化为中央液泡。我们的研究表明，在萌发的子叶中，质子泵和VM 23是新合成的，并以功能形式积累在蛋白体膜上。

We have investigated the membrane transport systems of plant vacuole. The vacuole performs numerous functions vital to cellular homeostasis, including the accumulation of metabolites and ions. These vacuolar functions are supported by the primary and secondary active transport systems. Two proton pumps, namely H^+-ATPase and H^+-pyrophosphatase, acidify the vacuole and create a membrane potential, which is utilized as an energy source for the secondary transport systems. In addition to the two proton pumps, we had analyzed a very hydrophobic protein, tentatively named VM23, which is thought to be a water channel of vacuolar membrane.In this study, we obtained the following findings. (1) H^+-pyrophosphatase widely distributes among green plants including fern and moss. Thus, H^+-pyrophosphatase is a common enzyme of green plant. However, VM23 were not found in the vacuolar membranes from Acetabularia, Chara, Conocephalum, or Kalanchoe. (2) We got a cDNA for barley H^+-pyrophosphatase. In the primary amino acid sequence estimated from the cDNA,we found two important parts, a catalytic part for substrate hydrolysis and a proton-channel forming domain. This information supports our proposal that a single polypeptide of 73 kilodaltons not only hydrolyses inorganic pyrophosphate but also transports protons across the membrane. (3) It is known that protein body transforms to central vacuole in pumpkin cotyledon during germination. Our investigation revealed that in the germinating cotyledons the proton pumps and VM23 are newly synthesized and accumulated in the protein-body membrane as functional forms.