背景:甘蔗糖蜜,富含蔗糖,葡萄糖,和果糖,由于其丰富和低成本,为工业发酵提供了有前途的碳源。然而,挑战来自同时利用多种糖和碳分解代谢物抑制(CCR)。尽管它的营养成分,大肠杆菌中的蔗糖代谢,除了W菌株,仍然知之甚少,阻碍其在微生物发酵中的使用。在这项研究中,大肠杆菌W被设计为提高糖消耗率并克服CCR。这是通过整合合成设计的csc操纵子和优化葡萄糖和果糖共利用途径来实现的。这些进步有助于有效利用甘蔗糖蜜生产3-羟基丙酸(3-HP),促进可持续的生化生产过程。
结果:在这项研究中,我们解决了与大肠杆菌W中糖代谢相关的挑战,重点提高蔗糖消耗和提高葡萄糖-果糖的共利用。通过对蔗糖利用系统的针对性工程,我们通过调节csc操纵子组件的表达来实现加速的蔗糖消耗率,CSCB,CSCK,cscA,和CSCR。我们的发现揭示了csc基因的单顺反子表达与cscR的缺失,导致最佳的蔗糖利用率,而没有显著的生长负担。此外,我们通过调节FruR与果糖PTS调节子的结合动力学,成功地减轻了果糖分解代谢物的抑制,实现葡萄糖和果糖的几乎等效的共同利用。为了验证我们工程菌株的工业适用性,我们追求3-HP生产甘蔗糖蜜。通过整合异源基因和优化代谢途径,与以前的研究相比,我们在3-HP滴度方面取得了改善.此外,甘油醛-3-磷酸脱氢酶(gapA)抑制有助于碳通量的重新分布,增强糖蜜转化为3-HP。
结论:尽管蔗糖代谢受到限制,重新设计的大肠杆菌W菌株,擅长利用甘蔗糖蜜,是工业发酵的宝贵资产。其合成的csc操纵子增加蔗糖消耗,同时减轻CCR提高葡萄糖-果糖的共同利用。这些增强,再加上对gapA的压制,旨在有效地将甘蔗糖蜜转化为3-HP,解决工业应用中蔗糖和果糖代谢的局限性。
BACKGROUND: Sugarcane molasses, rich in sucrose, glucose, and
fructose, offers a promising carbon source for industrial fermentation due to its abundance and low cost. However, challenges arise from the simultaneous utilization of multiple sugars and carbon catabolite repression (CCR). Despite its nutritional content, sucrose metabolism in Escherichia coli, except for W strain, remains poorly understood, hindering its use in microbial fermentation. In this study, E. coli W was engineered to enhance sugar consumption rates and overcome CCR. This was achieved through the integration of a synthetically designed csc operon and the optimization of glucose and
fructose co-utilization pathways. These advancements facilitate efficient utilization of sugarcane molasses for the production of 3-hydroxypropionic acid (3-HP), contributing to sustainable biochemical production processes.
RESULTS: In this study, we addressed challenges associated with sugar metabolism in E. coli W, focusing on enhancing sucrose consumption and improving glucose-fructose co-utilization. Through targeted engineering of the sucrose utilization system, we achieved accelerated sucrose consumption rates by modulating the expression of the csc operon components, cscB, cscK, cscA, and cscR. Our findings revealed that monocistronic expression of the csc genes with the deletion of cscR, led to optimal sucrose utilization without significant growth burden. Furthermore, we successfully alleviated
fructose catabolite repression by modulating the binding dynamics of FruR with the
fructose PTS regulon, enabling near-equivalent co-utilization of glucose and
fructose. To validate the industrial applicability of our engineered strain, we pursued 3-HP production from sugarcane molasses. By integrating heterologous genes and optimizing metabolic pathways, we achieved improvements in 3-HP titers compared to previous studies. Additionally, glyceraldehyde-3-phosphate dehydrogenase (gapA) repression aids in carbon flux redistribution, enhancing molasses conversion to 3-HP.
CONCLUSIONS: Despite limitations in sucrose metabolism, the redesigned E. coli W strain, adept at utilizing sugarcane molasses, is a valuable asset for industrial fermentation. Its synthetic csc operon enhances sucrose consumption, while mitigating CCR improves glucose-
fructose co-utilization. These enhancements, coupled with repression of gapA, aim to efficiently convert sugarcane molasses into 3-HP, addressing limitations in sucrose and fructose metabolism for industrial applications.