Keegan F, Blum J J
Department of Cell Biology, Duke University Medical Center, Durham, NC 27710.
Mol Biochem Parasitol. 1990 Mar;39(2):235-45. doi: 10.1016/0166-6851(90)90062-q.
Leishmania major promastigotes grown in late log phase were incubated with glucose as sole exogenous carbon source in the presence of 5% CO2 and the amounts of glucose consumed and of the major products formed--succinate, pyruvate, alanine, acetate, glycerol, and D-lactate--were measured as a function of pO2. Glucose consumption increased as pO2 was lowered to 6% (a positive Pasteur effect) and then declined to the same level at 95% N2 as at 95% O2. The production of D-lactate and of glycerol increased as pO2 dropped from 95%, reaching a maximum at about 2% O2. Succinate production, however, increased dramatically when pO2 was reduced to 6% and remained at that level with further reduction of pO2. The amount of succinate produced relative to the amount of glucose carbon consumed suggests utilization of an endogenous carbon source. Acetate production did not change between 95% O2 and 6% O2 and then declined with decreasing pO2. These observations suggest the presence of two sensors, one with a high and one with a low affinity for oxygen. When glycerol or alanine were the only exogenous sources of carbon, the primary products released were acetate and succinate. Acetate production from alanine declined slightly as pO2 was reduced to 2%, and then dropped markedly when pO2 was reduced to 0%. Acetate production from glycerol increased over 4-fold when the pO2 was reduced from 95% to 4%, and then declined with further reduction in pO2. No succinate was formed from either substrate until complete anaerobiosis. This pattern of response, while differing from that when glucose was sole exogenous carbon source, is also consistent with the regulation of metabolism by a high and a low affinity O2 sensor. Cells from cultures in early stationary phase, before the appearance of metacyclic forms, consumed glucose at about the same rate as log phase promastigotes, but did not show a Pasteur effect. Stationary cells also consumed glycerol at the same rate as did log phase promastigotes, but consumed alanine at a much lower rate. Reduction of pO2 affected product formation from each of these substrates differently than for log phase promastigotes, demonstrating the sensitivity of several pathways of intermediary metabolism to regulation by pO2 during the transition from log to stationary phase.
将处于对数后期生长阶段的硕大利什曼原虫前鞭毛体，在5%二氧化碳存在的条件下，以葡萄糖作为唯一的外源碳源进行培养，并测定葡萄糖的消耗量以及形成的主要产物——琥珀酸、丙酮酸、丙氨酸、乙酸盐、甘油和D - 乳酸——随pO₂的变化情况。随着pO₂降至6%，葡萄糖消耗量增加（正巴斯德效应），然后在95%氮气环境下与95%氧气环境下降至相同水平。当pO₂从95%下降时，D - 乳酸和甘油的产量增加，在约2%氧气时达到最大值。然而，当pO₂降至6%时，琥珀酸产量急剧增加，并在pO₂进一步降低时保持在该水平。相对于消耗的葡萄糖碳量，产生的琥珀酸量表明利用了内源性碳源。在95%氧气和6%氧气之间，乙酸盐产量没有变化，然后随着pO₂降低而下降。这些观察结果表明存在两种传感器，一种对氧气具有高亲和力，另一种具有低亲和力。当甘油或丙氨酸是唯一的外源碳源时，释放的主要产物是乙酸盐和琥珀酸。当pO₂降至2%时，丙氨酸产生的乙酸盐略有下降，然后在pO₂降至0%时显著下降。当pO₂从95%降至4%时，甘油产生的乙酸盐增加超过4倍，然后随着pO₂进一步降低而下降。在完全厌氧之前，两种底物都不会形成琥珀酸。这种反应模式虽然与以葡萄糖作为唯一外源碳源时不同，但也与由高亲和力和低亲和力氧气传感器调节代谢相一致。在出现后循环形式之前，处于早期稳定期培养物中的细胞消耗葡萄糖的速率与对数期前鞭毛体大致相同，但没有显示出巴斯德效应。稳定期细胞消耗甘油的速率也与对数期前鞭毛体相同，但消耗丙氨酸的速率要低得多。与对数期前鞭毛体相比，pO₂的降低对这些底物中每种底物的产物形成影响不同，这表明在从对数期到稳定期的转变过程中，中间代谢的几种途径对pO₂调节敏感。
