Inui Hiroshi, Ishikawa Takahiro, Tamoi Masahiro
Department of Nutrition, Osaka Prefecture University, 30-7-3 Habikino, Habikino, Osaka, 583-8555, Japan.
Faculty of Life and Environmental Science, Department of Life Science and Biotechnology, Shimane University, 1060 Nishikawatsu, Matsue, Shimane, 690-8504, Japan.
Adv Exp Med Biol. 2017;979:269-283. doi: 10.1007/978-3-319-54910-1_13.
In Euglena cells under anaerobic conditions, paramylon, the storage polysaccharide, is promptly degraded and converted to wax esters. The wax esters synthesized are composed of saturated fatty acids and alcohols with chain lengths of 10-18, and the major constituents are myristic acid and myristyl alcohol. Since the anaerobic cells gain ATP through the conversion of paramylon to wax esters, the phenomenon is named "wax ester fermentation". The wax ester fermentation is quite unique in that the end products, i.e. wax esters, have relatively high molecular weights, are insoluble in water, and accumulate in the cells, in contrast to the common fermentation end products such as lactic acid and ethanol.A unique metabolic pathway involved in the wax ester fermentation is the mitochondrial fatty acid synthetic system. In this system, fatty acid are synthesized by the reversal of β-oxidation with an exception that trans-2-enoyl-CoA reductase functions instead of acyl-CoA dehydrogenase. Therefore, acetyl-CoA is directly used as a C donor in this fatty acid synthesis, and the conversion of acetyl-CoA to malonyl-CoA, which requires ATP, is not necessary. Consequently, the mitochondrial fatty acid synthetic system makes possible the net gain of ATP through the synthesis of wax esters from paramylon. In addition, acetyl-CoA is provided in the anaerobic cells from pyruvate by the action of a unique enzyme, oxygen sensitive pyruvate:NADP oxidoreductase, instead of the common pyruvate dehydrogenase multienzyme complex.Wax esters produced by anaerobic Euglena are promising biofuels because myristic acid (C) in contrast to other algal produced fatty acids, such as palmitic acid (C) and stearic acid (C), has a low freezing point making it suitable as a drop-in jet fuel. To improve wax ester production, the molecular mechanisms by which wax ester fermentation is regulated in response to aerobic and anaerobic conditions have been gradually elucidated by identifying individual genes related to the wax ester fermentation metabolic pathway and by comprehensive gene/protein expression analysis. In addition, expression of the cyanobacterial Calvin cycle fructose-1,6-bisphosphatase/sedohepturose-1,7-bisphosphatase, in Euglena provided photosynthesis resulting in increased paramylon accumulation enhancing wax ester production. This chapter will discuss the biochemistry of the wax ester fermentation, recent advances in our understanding of the regulation of the wax ester fermentation and genetic engineering approaches to increase production of wax esters for biofuels.
在厌氧条件下的眼虫细胞中，储存多糖副淀粉会迅速降解并转化为蜡酯。合成的蜡酯由链长为10 - 18的饱和脂肪酸和醇组成，主要成分是肉豆蔻酸和肉豆蔻醇。由于厌氧细胞通过将副淀粉转化为蜡酯来获取ATP，这种现象被称为“蜡酯发酵”。蜡酯发酵相当独特，因为其终产物蜡酯具有相对较高的分子量，不溶于水且在细胞中积累，这与乳酸和乙醇等常见发酵终产物不同。蜡酯发酵所涉及的独特代谢途径是线粒体脂肪酸合成系统。在这个系统中，脂肪酸通过β - 氧化的逆向反应合成，不同的是反式 - 2 - 烯酰辅酶A还原酶发挥作用而不是酰基辅酶A脱氢酶。因此，乙酰辅酶A在这种脂肪酸合成中直接用作碳源，不需要将乙酰辅酶A转化为丙二酰辅酶A（这需要ATP）。因此，线粒体脂肪酸合成系统通过从副淀粉合成蜡酯使得ATP的净增加成为可能。此外，厌氧细胞中的乙酰辅酶A由丙酮酸通过一种独特的酶——氧敏感的丙酮酸：NADP氧化还原酶的作用提供，而不是通过常见的丙酮酸脱氢酶多酶复合体。厌氧眼虫产生的蜡酯是很有前景的生物燃料，因为与其他藻类产生的脂肪酸如棕榈酸（C）和硬脂酸（C）相比，肉豆蔻酸（C）的冰点较低，使其适合作为直接替代喷气燃料。为了提高蜡酯产量，通过鉴定与蜡酯发酵代谢途径相关的单个基因以及进行全面的基因/蛋白质表达分析，蜡酯发酵在有氧和厌氧条件下如何被调控的分子机制已逐渐被阐明。此外，在眼虫中表达蓝藻卡尔文循环果糖 - 1,6 - 二磷酸酶/景天庚酮糖 - 1,7 - 二磷酸酶，可实现光合作用，导致副淀粉积累增加，从而提高蜡酯产量。本章将讨论蜡酯发酵的生物化学、我们对蜡酯发酵调控理解的最新进展以及增加用于生物燃料的蜡酯产量的基因工程方法。
