The key difference in the modified Embden–Meyerhof glycolytic pathway in hyperthermophilic Archaea, such as Pyrococcus furiosus, occurs at the conversion from glyceraldehyde-3-phosphate (GAP) to 3-phosphoglycerate (3-PG) where the typical intermediate 1,3-bisphosphoglycerate (1,3-BPG) is not present. The absence of the ATP-yielding step catalyzed by phosphoglycerate kinase (PGK) alters energy yield, redox energetics, and kinetics of carbohydrate metabolism. Either of the two enzymes, ferredoxin-dependent glyceraldehyde-3-phosphate ferredoxin oxidoreductase (GAPOR) or NADP⁺-dependent non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPN), responsible for this “bypass” reaction, could be deleted individually without impacting viability, albeit with differences in native fermentation product profiles. Furthermore, P. furiosus was viable in the gluconeogenic direction (growth on pyruvate or peptides plus elemental sulfur) in a ΔgapnΔgapor strain. Ethanol was utilized as a proxy for potential heterologous products (e.g., isopropanol, butanol, fatty acids) that require reducing equivalents (e.g., NAD(P)H, reduced ferredoxin) generated from glycolysis. Insertion of a single gene encoding the thermostable NADPH-dependent primary alcohol dehydrogenase (adhA) (Tte_0696) from Caldanaerobacter subterraneus, resulted in a strain producing ethanol via the previously established aldehyde oxidoreductase (AOR) pathway. This strain demonstrated a high ratio of ethanol over acetate (> 8:1) at 80 °C and enabled ethanol production up to 85 °C, the highest temperature for bio-ethanol production reported to date.

中文翻译：

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激烈热球菌糖酵解途径的修饰及其对代谢工程的影响。

嗜热古生菌（如激烈热球菌）在修饰的Embden-Meyerhof糖酵解途径中的关键区别发生在从3-磷酸甘油醛（GAP）到3-磷酸甘油酸酯（3-PG）的转化处，其中典型的中间体1,3-不存在双磷酸甘油酸酯（1,3-BPG）。磷酸甘油酸激酶（PGK）催化的ATP生成步骤的缺失会改变能量产量，氧化还原能量和碳水化合物代谢的动力学。两种酶中的一种，铁氧还蛋白依赖性三磷酸甘油醛-磷酸铁氧还蛋白氧化还原酶（GAPOR）或NADP ⁺尽管存在天然发酵产物特性的差异，但可以单独删除导致该“旁路”反应的非依赖性非磷酸化甘油醛-3-磷酸甘油醛脱氢酶（GAPN），而不会影响生存力。此外，P.球菌是在糖异生方向（上丙酮酸或肽加元素硫生长）在一个可行的ΔgapnΔgapor菌株。乙醇被用作潜在异源产品（例如，异丙醇，丁醇，脂肪酸）的代用品，这些产品需要从糖酵解中产生的还原当量（例如，NAD（P）H，还原铁氧还蛋白）。单个基因插入编码热稳定的NADPH依赖性伯醇脱氢酶（ADHA从）（Tte_0696）Caldanaerobacter subterraneus导致通过先前建立的醛氧化还原酶（AOR）途径生产乙醇的菌株。该菌株在80°C时显示出乙醇与乙酸盐的比率高（> 8：1），并使乙醇的生产温度高达85°C，这是迄今为止报道的生物乙醇生产的最高温度。