Engineering Escherichia coli Nicotinic Acid Mononucleotide Adenylyltransferase for Fully Active Amidated NAD Biosynthesis

doi:10.1128/AEM.00692-17

	Engineering Escherichia coli Nicotinic Acid Mononucleotide Adenylyltransferase for Fully Active Amidated NAD Biosynthesis
	Wang, Xueying 2,3; Zhou, Yongjin J.2; Wang, Lei 2; Liu, Wujun 2; Liu, Yuxue 2,3; Peng, Chang 2,3; Zhao, Zongbao K.1,2
刊名	APPLIED AND ENVIRONMENTAL MICROBIOLOGY
	2017-07-01
卷号	83
关键词	Cofactor Engineering Nad Biosynthesis Protein Engineering
DOI	10.1128/AEM.00692-17
英文摘要	NAD and its reduced form NADH function as essential redox cofactors and have major roles in determining cellular metabolic features. NAD can be synthesized through the deamidated and amidated pathways, for which the key reaction involves adenylylation of nicotinic acid mononucleotide (NaMN) and nicotinamide mononucleotide (NMN), respectively. In Escherichia coli, NAD de novo biosynthesis depends on the protein NadD-catalyzed adenylylation of NaMN to nicotinic acid adenine dinucleotide (NaAD), followed by NAD synthase-catalyzed amidation. In this study, we engineered NadD to favor NMN for improved amidated pathway activity. We designed NadD mutant libraries, screened by a malic enzyme-coupled colorimetric assay, and identified two variants, 11B4 (Y84V/Y118D) and 16D8 (A86W/Y118N), with a high preference for NMN. Whereas in the presence of NMN both variants were capable of enabling the viability of cells of E. coli BW25113-derived NAD-auxotrophic strain YJE003, for which the last step of the deamidated pathway is blocked, the 16D8 expression strain could grow without exogenous NMN and accumulated a higher cellular NAD(H) level than BW25113 in the stationary phase. These mutants established fully active amidated NAD biosynthesis and offered a new opportunity to manipulate NAD metabolism for biocatalysis and metabolic engineering.
语种	英语
WOS记录号	WOS:000403495700023
内容类型	期刊论文
源URL	[http://cas-ir.dicp.ac.cn/handle/321008/152103]
专题	大连化学物理研究所_中国科学院大连化学物理研究所
作者单位	1.Chinese Acad Sci, Dalian Inst Chem Phys, State Key Lab Catalysis, Dalian, Peoples R China 2.Chinese Acad Sci, Dalian Inst Chem Phys, Div Biotechnol, Dalian, Peoples R China 3.Univ Chinese Acad Sci, Beijing, Peoples R China
推荐引用方式 GB/T 7714	Wang, Xueying,Zhou, Yongjin J.,Wang, Lei,et al. Engineering Escherichia coli Nicotinic Acid Mononucleotide Adenylyltransferase for Fully Active Amidated NAD Biosynthesis[J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY,2017,83.
APA	Wang, Xueying.,Zhou, Yongjin J..,Wang, Lei.,Liu, Wujun.,Liu, Yuxue.,...&Zhao, Zongbao K..(2017).Engineering Escherichia coli Nicotinic Acid Mononucleotide Adenylyltransferase for Fully Active Amidated NAD Biosynthesis.APPLIED AND ENVIRONMENTAL MICROBIOLOGY,83.
MLA	Wang, Xueying,et al."Engineering Escherichia coli Nicotinic Acid Mononucleotide Adenylyltransferase for Fully Active Amidated NAD Biosynthesis".APPLIED AND ENVIRONMENTAL MICROBIOLOGY 83(2017).