革兰氏阳性模型细菌枯草芽孢杆菌用于许多生物技术应用,包括大规模生产维生素。对于维生素B5,辅酶A合成的前体,到目前为止还没有成熟的发酵过程,涉及这种维生素的代谢途径只有部分了解。在这项研究中,我们已经阐明了枯草芽孢杆菌中泛酸和辅酶A的生物合成的完整途径。泛酸不仅可以合成,而且可以从培养基中提取。我们已经鉴定了参与泛酸生物合成和摄取的酶和转运蛋白。枯草芽孢杆菌中的高亲和力维生素B5摄取需要ATP驱动的能量耦合因子转运蛋白,其中PanU(以前是YhfU)作为底物特异性亚基。此外,我们已经确定了即使在没有泛酸合成的情况下也作用于复杂培养基的辅酶A获取的补救途径。该途径需要重新连接硫代谢,导致半胱氨酸转运蛋白的表达增加。在救助途径中,细菌进口半胱氨酸,一种新的天然代谢产物,使用胱氨酸运输系统TcyJKLMN。这项工作为开发使用枯草芽孢杆菌生产维生素B5和辅酶A的有效方法奠定了基础。
目的:维生素是动物和人类饮食的重要组成部分。因此,维生素是生物技术生产的重要目标。虽然已经为几种维生素开发了有效的发酵工艺,维生素B5(泛酸)的情况并非如此,辅酶A的前体。我们已经阐明了生物技术主力枯草芽孢杆菌中辅酶A生物合成的完整途径。此外,在这项研究中发现了辅酶A合成的补救途径。通常,这个途径依赖于pantetheine;然而,我们在缺乏泛酸生物合成途径的突变体中观察到了在复杂培养基上补救途径的活性,即使没有补充泛酸。由于获得影响半胱氨酸代谢调节的突变,这需要通过表达胱氨酸转运蛋白来重新连接代谢。这表明隐藏的“地下代谢”如何导致新的代谢途径的快速形成。
The Gram-positive model bacterium Bacillus subtilis is used for many biotechnological applications, including the large-scale production of vitamins. For vitamin B5, a precursor for coenzyme A synthesis, there is so far no established fermentation process available, and the metabolic pathways that involve this vitamin are only partially understood. In this study, we have elucidated the complete pathways for the biosynthesis of pantothenate and coenzyme A in B. subtilis. Pantothenate can not only be synthesized but also be taken up from the medium. We have identified the enzymes and the transporter involved in the pantothenate biosynthesis and uptake. High-affinity vitamin B5 uptake in B. subtilis requires an ATP-driven energy coupling factor transporter with PanU (previously YhfU) as the substrate-specific subunit. Moreover, we have identified a salvage pathway for coenzyme A acquisition that acts on complex medium even in the absence of pantothenate synthesis. This pathway requires rewiring of sulfur metabolism resulting in the increased expression of a cysteine transporter. In the salvage pathway, the bacteria import cysteinopantetheine, a novel naturally occurring metabolite, using the cystine transport system TcyJKLMN. This work lays the foundation for the development of effective processes for vitamin B5 and coenzyme A production using B. subtilis.
OBJECTIVE: Vitamins are essential components of the diet of animals and humans. Vitamins are thus important targets for biotechnological production. While efficient fermentation processes have been developed for several vitamins, this is not the case for vitamin B5 (pantothenate), the precursor of coenzyme A. We have elucidated the complete pathway for coenzyme A biosynthesis in the biotechnological workhorse Bacillus subtilis. Moreover, a salvage pathway for coenzyme A synthesis was found in this study. Normally, this pathway depends on pantetheine; however, we observed activity of the salvage pathway on complex medium in mutants lacking the pantothenate biosynthesis pathway even in the absence of supplemented pantetheine. This required rewiring of metabolism by expressing a cystine transporter due to acquisition of mutations affecting the regulation of cysteine metabolism. This shows how the hidden \"underground metabolism\" can give rise to the rapid formation of novel metabolic pathways.