GIGA-Neurosciences (B36), University of Liège, Avenue de l'Hôpital 1, B-4000 Liège 1 (Sart Tilman), Belgium B-4000 Liège, Belgium.
J Biol Chem. 2010 Jan 1;285(1):583-94. doi: 10.1074/jbc.M109.054379. Epub 2009 Nov 11.
In animals, thiamine deficiency leads to specific brain lesions, generally attributed to decreased levels of thiamine diphosphate, an essential cofactor in brain energy metabolism. However, another far less abundant derivative, thiamine triphosphate (ThTP), may also have a neuronal function. Here, we show that in the rat brain, ThTP is essentially present and synthesized in mitochondria. In mitochondrial preparations from brain (but not liver), ThTP can be produced from thiamine diphosphate and P(i). This endergonic process is coupled to the oxidation of succinate or NADH through the respiratory chain but cannot be energized by ATP hydrolysis. ThTP synthesis is strongly inhibited by respiratory chain inhibitors, such as myxothiazol and inhibitors of the H(+) channel of F(0)F(1)-ATPase. It is also impaired by disruption of the mitochondria or by depolarization of the inner membrane (by protonophores or valinomycin), indicating that a proton-motive force (Deltap) is required. Collapsing Deltap after ThTP synthesis causes its rapid disappearance, suggesting that both synthesis and hydrolysis are catalyzed by a reversible H(+)-translocating ThTP synthase. The synthesized ThTP can be released from mitochondria in the presence of external P(i). However, ThTP probably does not accumulate in the cytoplasm in vivo, because it is not detected in the cytosolic fraction obtained from a brain homogenate. Our results show for the first time that a high energy triphosphate compound other than ATP can be produced by a chemiosmotic type of mechanism. This might shed a new light on our understanding of the mechanisms of thiamine deficiency-induced brain lesions.
在动物中，硫胺素缺乏会导致特定的脑损伤，通常归因于硫胺素二磷酸（一种脑能量代谢必需的辅酶）水平降低。然而，另一种含量少得多的衍生物，三磷酸硫胺素（ThTP），也可能具有神经元功能。在这里，我们表明在大鼠脑中，ThTP 主要存在于线粒体中并由其合成。在线粒体制剂中（但不在肝组织中），ThTP 可以由硫胺素二磷酸和 P(i) 产生。这个吸能过程通过呼吸链与琥珀酸或 NADH 的氧化偶联，但不能通过 ATP 水解来供能。ThTP 的合成强烈受到呼吸链抑制剂（如鱼藤酮和 F(0)F(1)-ATP 酶 H(+)通道抑制剂）的抑制。它还受到线粒体破坏或内膜去极化（通过质子载体或缬氨霉素）的影响，表明需要质子动势（Deltap）。ThTP 合成后 Deltap 的崩溃导致其迅速消失，这表明合成和水解都由可逆的 H(+)-转运 ThTP 合酶催化。在存在外部 P(i)的情况下，合成的 ThTP 可以从线粒体中释放出来。然而，ThTP 可能不会在体内的细胞质中积累，因为在从脑匀浆获得的胞质部分中未检测到它。我们的结果首次表明，一种非 ATP 的高能三磷酸化合物可以通过一种化学渗透型机制产生。这可能为我们理解硫胺素缺乏引起的脑损伤机制提供新的视角。
