Pablos Tania E, Sigala Juan Carlos, Le Borgne Sylvie, Lara Alvaro R
Departamento de Procesos y Tecnología, Universidad Autónoma Metropolitana-Cuajimalpa, México; Doctorado en Ciencias Biológicas y de la Salud de la Universidad Autónoma Metropolitana, México.
Biotechnol J. 2014 Jun;9(6):791-9. doi: 10.1002/biot.201300388. Epub 2014 Mar 18.
Overflow metabolism is a prevalent problem for aerobic cultivations of Escherichia coli. Although several process and molecular approaches have been applied to prevent overflow metabolism, these approaches often result in reductions in growth rate, biomass yield or accumulation of other byproducts. In this report, we present an alternative approach based on increasing the efficiency of aerobic metabolism by the expression of the Vitreoscilla stercoraria hemoglobin (VHb) to avoid overflow metabolism. VHb is expected to increase the consumption of NADH in the respiratory chain, leading to increased activity of the tricarboxylic acid (TCA) cycle. This would result in a faster consumption of acetyl Co-A and a decrease in acetate production. When this strategy was tested in E. coli strains, acetate production decreased by 50% in MG1655 and more than 90% in W3110, without affecting growth rates or biomass yields. VHb expression in mutant strains with higher TCA activity and reduced acetate formation resulted in a significant increase in growth and glucose consumption rates, whereas acetate production did not increase. The results presented here show that enhancing the efficiency of aerobic metabolism is a valuable approach to avoid overflow metabolism in E. coli and to attain high cell densities in batch mode.
溢流代谢是大肠杆菌好氧培养中普遍存在的问题。尽管已经应用了几种工艺和分子方法来防止溢流代谢，但这些方法往往会导致生长速率、生物量产量降低或其他副产物的积累。在本报告中，我们提出了一种基于表达嗜尸生玻瓶菌血红蛋白（VHb）来提高好氧代谢效率以避免溢流代谢的替代方法。预计VHb会增加呼吸链中NADH的消耗，从而导致三羧酸（TCA）循环活性增强。这将导致乙酰辅酶A消耗加快，乙酸盐产量降低。当在大肠杆菌菌株中测试该策略时，MG1655中的乙酸盐产量降低了50%，W3110中的乙酸盐产量降低了90%以上，且不影响生长速率或生物量产量。在具有较高TCA活性和减少乙酸盐形成的突变菌株中表达VHb导致生长和葡萄糖消耗速率显著增加，而乙酸盐产量并未增加。此处给出的结果表明，提高好氧代谢效率是避免大肠杆菌溢流代谢并在分批模式下实现高细胞密度的一种有价值的方法。
