Garlick Andrew P, Moore Catherine, Kruger Nicholas J
Department of Plant Sciences, University of Oxford, South Parks Road, Oxford, OX1 3RB, UK.
Planta. 2002 Dec;216(2):265-72. doi: 10.1007/s00425-002-0842-1. Epub 2002 Aug 10.
The aim of this work was to examine the metabolism of exogenous gluconate by a 4-day-old cell suspension culture of Arabidopsis thaliana (L.) Heynh. Release of (14)CO(2) from [1-(14)C]gluconate was dependent on the concentration in the medium and could be resolved into a substrate-saturable component (apparent K(m) of approximately 0.4 mM) and an unsaturable component. At an external concentration of 0.3 mM, the rate of decarboxylation of applied gluconate was 0.2% of the rate of oxygen consumption by the cells. There was no effect of 0.3 mM gluconate on the rate of oxygen consumption, or on the rate of (14)CO(2) release from either [1-(14)C]glucose or [6-(14)C]glucose by the culture. The following observations argue that gluconate taken up by the cells is metabolised by direct phosphorylation to 6-phosphogluconate and subsequent decarboxylation through 6-phosphogluconate dehydrogenase. First, more than 95% of the label released from [1-(14)C]gluconate during metabolism by the cell culture was recovered as (14)CO(2). Secondly, inhibition of the oxidative pentose phosphate pathway (OPPP) by treatment with 6-aminonicotinamide preferentially inhibited release of (14)CO(2) from [1-(14)C]gluconate relative to that from [1-(14)C]glucose. Thirdly, perturbation of glucose metabolism by glucosamine did not affect (14)CO(2) from [1-(14)C]gluconate. Fourth, stimulation of the OPPP by phenazine methosulphate stimulated release of (14)CO(2) from [1-(14)C]gluconate to a far greater extent than that from [1-(14)C]glucose. It is proposed that measurement of (14)CO(2) from [1-(14)C]gluconate provides a simple and sensitive technique for monitoring flux through the OPPP pathway in plants.
这项工作的目的是研究拟南芥（L.）海因茨4日龄细胞悬浮培养物对外源葡萄糖酸盐的代谢情况。[1-(14)C]葡萄糖酸盐中(14)CO(2)的释放取决于培养基中的浓度，并且可以分解为底物饱和成分（表观K(m)约为0.4 mM）和不饱和成分。在外部浓度为0.3 mM时，施加的葡萄糖酸盐的脱羧速率是细胞耗氧速率的0.2%。0.3 mM葡萄糖酸盐对耗氧速率或培养物从[1-(14)C]葡萄糖或[6-(14)C]葡萄糖释放(14)CO(2)的速率没有影响。以下观察结果表明，细胞摄取的葡萄糖酸盐通过直接磷酸化代谢为6-磷酸葡萄糖酸盐，随后通过6-磷酸葡萄糖酸脱氢酶进行脱羧。首先，在细胞培养物代谢[1-(14)C]葡萄糖酸盐过程中释放的标记物中，超过95%以(14)CO(2)的形式回收。其次，用6-氨基烟酰胺处理抑制氧化戊糖磷酸途径（OPPP），相对于从[1-(14)C]葡萄糖释放的(14)CO(2)，优先抑制[1-(14)C]葡萄糖酸盐释放(14)CO(2)。第三，氨基葡萄糖对葡萄糖代谢的干扰不影响[1-(14)C]葡萄糖酸盐释放(14)CO(2)。第四，吩嗪硫酸甲酯对OPPP的刺激对[1-(14)C]葡萄糖酸盐释放(14)CO(2)的刺激程度远大于对[1-(14)C]葡萄糖的刺激。有人提出，测量[1-(14)C]葡萄糖酸盐释放的(14)CO(2)为监测植物中通过OPPP途径的通量提供了一种简单而灵敏的技术。
