Key Laboratory of Bio-resources and Eco-environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu, China.
J Appl Microbiol. 2014 Jul;117(1):126-38. doi: 10.1111/jam.12510. Epub 2014 May 15.
To contribute to the improvement of methods for the regulation and production of higher alcohols using micro-organisms, we assessed the yields achieved using 10 decarboxylase genes from three different yeast species (Saccharomyces cerevisiae, Candida tropicalis and Pichia pastoris) by cloning them into vectors and overexpressing them in Escherichia coli hosts of different genotypes. Genes that produced the greatest yields in higher alcohol production were further assessed for the catalytic effects of the decarboxylase enzymes in the different E. coli hosts.
A major metabolic pathway is structured via overexpressing a series of five genes, to detect the effect of decarboxylase on the production of higher alcohols. Results suggested that these genes can facilitate production of specific types of higher alcohols by diverse types of E. coli. We also showed that they play direct roles in the metabolic pathways that lead to production of higher alcohols in E. coli. The gene ARO10 from S. cerevisiae produced the highest yields for producing isobutanol and isopentanol in the host JM109. Significant differences were found in the types of higher alcohols and yields produced within the same host, for the genes PAD1, GAD1, SPE1 from S. cerevisiae. Similar results were observed for the genes ODC1 and gadB from Candida tropicalis and P. pastoris, respectively.
Investigation of these genes for identification of the key enzymatic steps or regulatory pathways involved in the Ehrlich metabolic network to produce higher alcohols is paramount for producing biofuels. The selected genes are promising targets for the development of improved production strains.
This is the first published assessment of the effects of decarboxylases from different yeast species that were expressed in E. coli, for the production of higher alcohols. Our results provide guidance for future studies about the use of yeast enzymes for transforming or constructing a new metabolic pathway utilizing E. coli for the production of target higher alcohols.
为了改进利用微生物调节和生产高级醇的方法，我们评估了来自 3 种不同酵母（酿酒酵母、热带假丝酵母和巴斯德毕赤酵母）的 10 种脱羧酶基因在不同基因型大肠杆菌宿主中克隆到载体并过表达时的产量。进一步评估了在不同大肠杆菌宿主中产生高级醇产量最高的基因的脱羧酶的催化作用。
通过过表达一系列 5 个基因构建了一条主要的代谢途径，以检测脱羧酶对高级醇生产的影响。结果表明，这些基因可以促进不同类型的大肠杆菌生产特定类型的高级醇。我们还表明，它们在导致大肠杆菌中生产高级醇的代谢途径中发挥直接作用。来自酿酒酵母的 ARO10 基因在 JM109 宿主中产生异丁醇和异戊醇的产量最高。来自酿酒酵母的 PAD1、GAD1 和 SPE1 基因在同一宿主中产生的高级醇种类和产量存在显著差异。来自热带假丝酵母的 ODC1 和 gadB 基因以及来自巴斯德毕赤酵母的基因分别观察到类似的结果。
研究这些基因对于确定参与 Ehrlich 代谢网络生产高级醇的关键酶步骤或调控途径至关重要，这对于生产生物燃料至关重要。所选基因是开发改良生产菌株的有前途的目标。
这是首次评估不同酵母物种的脱羧酶在大肠杆菌中表达对高级醇生产的影响。我们的研究结果为利用酵母酶转化或构建利用大肠杆菌生产目标高级醇的新代谢途径的未来研究提供了指导。
