Metabolic engineering of low-pH-tolerant non-model yeast, Issatchenkia orientalis, for production of citramalate

IF 3.7 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Zong-Yen Wu , Wan Sun , Yihui Shen , Jimmy Pratas , Patrick F. Suthers , Ping-Hung Hsieh , Sudharsan Dwaraknath , Joshua D. Rabinowitz , Costas D. Maranas , Zengyi Shao , Yasuo Yoshikuni
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引用次数: 2

Abstract

Methyl methacrylate (MMA) is an important petrochemical with many applications. However, its manufacture has a large environmental footprint. Combined biological and chemical synthesis (semisynthesis) may be a promising alternative to reduce both cost and environmental impact, but strains that can produce the MMA precursor (citramalate) at low pH are required. A non-conventional yeast, Issatchenkia orientalis, may prove ideal, as it can survive extremely low pH. Here, we demonstrate the engineering of I. orientalis for citramalate production. Using sequence similarity network analysis and subsequent DNA synthesis, we selected a more active citramalate synthase gene (cimA) variant for expression in I. orientalis. We then adapted a piggyBac transposon system for I. orientalis that allowed us to simultaneously explore the effects of different cimA gene copy numbers and integration locations. A batch fermentation showed the genome-integrated-cimA strains produced 2.0 g/L citramalate in 48 h and a yield of up to 7% mol citramalate/mol consumed glucose. These results demonstrate the potential of I. orientalis as a chassis for citramalate production.

用于生产柠檬酸盐的低pH耐受性非模式酵母Issatchenkia orientalis的代谢工程
甲基丙烯酸甲酯(MMA)是一种重要的石油化工产品,有着广泛的应用。然而,它的制造有很大的环境足迹。生物和化学合成(半合成)可能是降低成本和环境影响的一种很有前途的替代方法,但需要能够在低pH下生产MMA前体(柠檬酸盐)的菌株。非传统酵母Issatchenkia orientalis可能被证明是理想的,因为它可以在极低的pH值下存活。在这里,我们展示了I.orientalis用于柠檬酸盐生产的工程。通过序列相似性网络分析和随后的DNA合成,我们选择了一种更具活性的柠檬酸合成酶基因(cimA)变体在东方I.orientalis中表达。然后,我们将piggyBac转座子系统用于东方I.orientalis,使我们能够同时探索不同cimA基因拷贝数和整合位置的影响。分批发酵表明,基因组整合的cimA菌株在48小时内产生2.0g/L的柠檬酸盐,并且产量高达7%摩尔柠檬酸盐/mol消耗的葡萄糖。这些结果证明了东方乳杆菌作为柠檬酸盐生产底盘的潜力。
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来源期刊
Metabolic Engineering Communications
Metabolic Engineering Communications Medicine-Endocrinology, Diabetes and Metabolism
CiteScore
13.30
自引率
1.90%
发文量
22
审稿时长
18 weeks
期刊介绍: Metabolic Engineering Communications, a companion title to Metabolic Engineering (MBE), is devoted to publishing original research in the areas of metabolic engineering, synthetic biology, computational biology and systems biology for problems related to metabolism and the engineering of metabolism for the production of fuels, chemicals, and pharmaceuticals. The journal will carry articles on the design, construction, and analysis of biological systems ranging from pathway components to biological complexes and genomes (including genomic, analytical and bioinformatics methods) in suitable host cells to allow them to produce novel compounds of industrial and medical interest. Demonstrations of regulatory designs and synthetic circuits that alter the performance of biochemical pathways and cellular processes will also be presented. Metabolic Engineering Communications complements MBE by publishing articles that are either shorter than those published in the full journal, or which describe key elements of larger metabolic engineering efforts.
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