Metabolic engineering of Cupriavidus necator H16 for improved chemoautotrophic growth and PHB production under oxygen-limiting conditions

doi:10.1016/j.ymben.2020.04.009

	Metabolic engineering of Cupriavidus necator H16 for improved chemoautotrophic growth and PHB production under oxygen-limiting conditions
	Tang, Ruohao 1,2; Weng, Caihong 1,2; Peng, Xiaowei 2; Han, Yejun 2
刊名	METABOLIC ENGINEERING
	2020-09-01
卷号	61 页码:11-23
关键词	Cupriavidus necator H16 Carbon dioxide Oxygen-limiting conditions Transcription Metabolic engineering Ployhydroxybutyrate
ISSN号	1096-7176
DOI	10.1016/j.ymben.2020.04.009
英文摘要	The oxygen-limiting condition promotes the accumulation of ployhydroxybutyrate (PHB) in C. necator H16, while the growth of which is restricted. Under autotrophic culture using carbon dioxide, hydrogen, and oxygen as substrates, the oxygen concentration below 6.9% (v/v) in the mixture is considered as a safe condition. It also expected to achieve cell rapid growth and large accumulation of PHB simultaneously under the oxygen-limiting condition in C. necator H16. In this study, a metabolically engineered strain capable of both rapid growth and large accumulation of PHB under oxygen-limiting conditions was constructed based on the transcriptomic analysis. In the comparative transcriptomic analysis, the genes related to energy-generating of C. necator H16 at autotrophic culture were downregulated under oxygen-limiting conditions (3%, v/v). Besides, the genes related to the key intermediates (pyruvate and acetyl-CoA) metabolism in PHB biosynthetic pathway were analyzed. Most of which were downregulated, except the genes ldh, iclA, and ackA2 respectively encoding L-lactate dehydrogenase, isocitrate lyase, and acetate kinase were upregulated under oxygen-limiting conditions (3%, v/v). The Vitreoscilla hemoglobin (VHb) has the ability to promote aerobic metabolism and energy generation. To promote the bacterium growth and improve the energy generation in C. necator H16 under oxygen-limiting conditions, the VHb gene was introduced into C. necator H16 with the optimized promoter PphaC1-j5. Moreover, VHb was localized to the periplasmic space of the bacterium by the traction of membrane-bound hydrogenase (MBH) signal peptide. By optimizing the knockout of different genes, it was found that knockout of ldh can improve PHB production and reduce the by-products. Finally, a recombinant strain Reh01 (p2M-pj-v) was constructed by heterologous expression of vgb and ldh knockout in C. necator H16. Compared with the control (Reh (p2)) under oxygen-limiting conditions (3%, v/v), the dry cell weight (DCW), PHB content, and PHB production of Reh01 (p2M-pj-v) increased by 31.0%, 30.9%, and 71.5%, respectively. From the perspectives of transcriptome and metabolic engineering, the work provides new ideas to achieve rapid cell growth and large PHB accumulation in C. necator under oxygen-limiting and autotrophic conditions.
资助项目	National Natural Science Foundation of China[21676279] ; National Key Research and Development Project of China[2019YFB1503800]
WOS关键词	VITREOSCILLA HEMOGLOBIN ; GENE-EXPRESSION ; HYDROGEN BACTERIA ; TANDEM PROMOTERS ; CELL-DENSITY ; CO2 ; CULTIVATION ; EXCRETION ; CULTURE ; O-2
WOS研究方向	Biotechnology & Applied Microbiology
语种	英语
出版者	ACADEMIC PRESS INC ELSEVIER SCIENCE
WOS记录号	WOS:000570475100002
资助机构	National Natural Science Foundation of China ; National Key Research and Development Project of China
内容类型	期刊论文
源URL	[http://ir.ipe.ac.cn/handle/122111/42161]
专题	中国科学院过程工程研究所
通讯作者	Peng, Xiaowei; Han, Yejun
作者单位	1.Univ Chinese Acad Sci, Sch Chem Engn, Beijing 100049, Peoples R China 2.Chinese Acad Sci, Inst Proc Engn, Natl Key Lab Biochem Engn, Beijing 100190, Peoples R China
推荐引用方式 GB/T 7714	Tang, Ruohao,Weng, Caihong,Peng, Xiaowei,et al. Metabolic engineering of Cupriavidus necator H16 for improved chemoautotrophic growth and PHB production under oxygen-limiting conditions[J]. METABOLIC ENGINEERING,2020,61:11-23.
APA	Tang, Ruohao,Weng, Caihong,Peng, Xiaowei,&Han, Yejun.(2020).Metabolic engineering of Cupriavidus necator H16 for improved chemoautotrophic growth and PHB production under oxygen-limiting conditions.METABOLIC ENGINEERING,61,11-23.
MLA	Tang, Ruohao,et al."Metabolic engineering of Cupriavidus necator H16 for improved chemoautotrophic growth and PHB production under oxygen-limiting conditions".METABOLIC ENGINEERING 61(2020):11-23.