University of Science and Technology of China, Hefei 230026, P.R. China.
Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, P.R. China.
J Microbiol Biotechnol. 2019 Aug 28;29(8):1288-1298. doi: 10.4014/jmb.1904.04019.
Bacterial ATP synthases drive ATP synthesis by a rotary mechanism, and play a vital role in physiology and cell metabolism. is well known as an industrial workhorse for amino acid production, and its ATP synthase operon contains eight structural genes and two adjacent genes, and . So far, the physiological functions of Cg1360 (GenBank CAF19908) and Cg1361 (GenBank CAF19909) remain unclear. Here, we showed that Cg1360 was a hydrophobic protein with four transmembrane helices (TMHs), while no TMH was found in Cg1361. Deletion of , but not , led to significantly reduced cell growth using glucose and acetic acid as carbon sources, reduced F1 portions in the membrane, reduced ATP-driven proton-pumping activity and ATPase activity, suggesting that Cg1360 plays an important role in ATP synthase function. The intracellular ATP concentration in the mutant was decreased to 72% of the wild type, while the NADH and NADPH levels in the mutant were increased by 29% and 26%, respectively. However, the mutant exhibited comparable intracellular ATP, NADH and NADPH levels with the wild-type strain. Moreover, the effect of deletion on L-valine production was examined in the L-valine-producing V-10 strain. The final production of L-valine in the V-10- mutant reached 9.2 ± 0.3 g/l in shake flasks, which was 14% higher than that of the V-10 strain. Thus, Cg1360 can be used as an effective engineering target by altering energy metabolism for the enhancement of amino acid production in .
细菌 ATP 合酶通过旋转机制驱动 ATP 合成，在生理和细胞代谢中起着至关重要的作用。 是生产氨基酸的工业主力，其 ATP 合酶操纵子包含 8 个结构基因和两个相邻基因 和 。到目前为止，Cg1360（GenBank CAF19908）和 Cg1361（GenBank CAF19909）的生理功能仍不清楚。在这里，我们表明 Cg1360 是一种具有四个跨膜螺旋（TMH）的疏水性蛋白，而 Cg1361 中没有 TMH。 缺失，但不是 ，导致以葡萄糖和乙酸为碳源时细胞生长显著减少，膜中 F1 部分减少，ATP 驱动的质子泵活性和 ATP 酶活性降低，表明 Cg1360 在 ATP 合酶功能中起重要作用。 突变体的细胞内 ATP 浓度降低到野生型的 72%，而 突变体的 NADH 和 NADPH 水平分别增加了 29%和 26%。然而， 突变体与野生型菌株的细胞内 ATP、NADH 和 NADPH 水平相当。此外，还在生产 L-缬氨酸的 V-10 菌株中检查了 缺失对 L-缬氨酸生产的影响。在摇瓶中，V-10- 突变体的 L-缬氨酸最终产量达到 9.2±0.3 g/L，比 V-10 菌株高 14%。因此，通过改变能量代谢，Cg1360 可以作为一种有效的工程靶点，用于增强 中的氨基酸生产。
