The monomer, 3-hydroxybutyrate (3HB), plays an interesting role as a precursor for antibiotics, vitamins, and bioplastics such as polyhydroxybutyrates (PHB). The biotechnological production of both compounds in Escherichia coli has seen increased interest in the last decade. The central metabolite in the 3HB production pathway is acetyl-CoA, the derivative of coenzyme A (CoA). Enriching the intracellular pool of these cofactors could improve 3HB titers. In our study, we opted to increase CoA titers by upregulating pantothenate kinase (PanK), the rate limiting step in CoA biosynthetic pathway. To this end, 4 PanKs genes of different taxonomic origins (mammalian, fungal, and bacterial) were individually expressed and evaluated in 3HB producing E. coli cells. In shake flask studies, strains expressing Aspergillus nidulans PankII and Mus musculus PanK1β achieved the highest 3HB titers. In a bioreactor fermentation, the strain harboring murine PanK1β resulted in 7.6 g/L compared to 5.4 g/L of 3HB in the control strain. Although structurally different from the bacterial PankI, our study showed that eukaryotic Panks can supplement the kinase activity in prokaryotes. Expressing the exogenous PanKs offer several advantages over the host’s enzyme; PanKII is only inhibited by acetyl-CoA, for which the 3HB-production system would provide a constant metabolic sink. Additionally, PanK1β is weakly regulated by acetyl-CoA, and its activity is stimulated by free CoA. Overexpressing eukaryotic PanKs constitutes a suitable strategy for improving 3HB titers in E. coli.

单体 3-羟基丁酸酯 (3HB) 作为抗生素、维生素和生物塑料（如聚羟基丁酸酯 (PHB)）的前体发挥着重要作用。在过去十年中，人们对大肠杆菌中这两种化合物的生物技术生产越来越感兴趣。3HB 生产途径中的中心代谢物是乙酰辅酶 A，辅酶 A (CoA) 的衍生物。丰富这些辅因子的细胞内库可以提高 3HB 滴度。在我们的研究中，我们选择通过上调泛酸激酶 (PanK)（CoA 生物合成途径中的限速步骤）来增加 CoA 滴度。为此，不同分类学起源（哺乳动物、真菌和细菌）的 4 个 PanKs 基因分别在产生 3HB 的大肠杆菌中表达和评估细胞。在摇瓶研究中，表达构巢曲霉PankII 和Mus musculus 的菌株PanK1β 达到了最高的 3HB 滴度。在生物反应器发酵中，携带鼠 PanK1β 的菌株产生 7.6 g/L，而对照菌株中的 3HB 为 5.4 g/L。尽管在结构上与细菌 PankI 不同，但我们的研究表明真核 Panks 可以补充原核生物中的激酶活性。表达外源性 PanKs 比宿主酶有几个优势；PanKII 仅受乙酰辅酶 A 抑制，为此 3HB 生产系统将提供恒定的代谢汇。此外，PanK1β 受乙酰辅酶 A 微弱调节，其活性受游离 CoA 刺激。过表达真核 PanKs 是提高大肠杆菌3HB 滴度的合适策略。