DICP OpenIR
Subject Area微生物学
Construction of E. coli NAD+ auxotrophic strains and the biotechnological application thereof
Zhou YJ(周雍进); Hou SH(侯淑华); Zhang SF(张素芳); Zhao ZB(赵宗保)
Source PublicationBook of Abstracts
Conference NameAsian Congress on Biotechnology
Conference Date2011-5-11
2011
Conference Place上海
Pages26-0
Publisher待补充
Publication Place待补充
Cooperation Status分会口头报告
Department1816
Funding Organization亚洲生物技术联合会
AbstractNAD+ and its reduced form NADH are essential cofactors in biological systems. They function as cofactors in over 300 redox reactions in vivo1. The level of NAD+ and NADH is tightly controlled by a variety of mechanisms including their biosynthesis and salvage. Therefore, it is difficult to answer some fundamental questions such as the minimal NAD+ level for cell growth and the biological consequences of abnormal activity of a specific NAD+-dependent enzyme. We expressed the NTT4 gene from Chlamydiae UWE25 in Escherichia coli BW25113, as the NTT4 protein was reported as a NAD+ transporter that can specifically transport intact NAD+ across cytoplasmic membrane2. We knocked out the nadC gene responsible for de novo biosynthesis of NAD+ and constructed the strain E. coli BW25113 (ΔnadC, NTT4). It was found that NAD+ in the culture media could significantly promote the growth of BW25113 (ΔnadC, NTT4), suggesting that the NTT4 protein was functional in E. coli (Fig A, B). We then disrupted the other two genes, nadD and nadE, and obtained the strains E. coli BW25113 (ΔnadD, NTT4) and BW25113 (ΔnadE, NTT4). Cell growth of these two strains are depending on exogenous NAD+ supplemented in the media, suggesting that we have successfully engineered E. coli to hold an NAD+ auxotrophic phenotype. We are carrying out a number of experiments using these NAD+ auxotrophic strains to address some interesting questions which may not be able to do otherwise. Results will be discussed during the conference.; NAD+ and its reduced form NADH are essential cofactors in biological systems. They function as cofactors in over 300 redox reactions in vivo1. The level of NAD+ and NADH is tightly controlled by a variety of mechanisms including their biosynthesis and salvage. Therefore, it is difficult to answer some fundamental questions such as the minimal NAD+ level for cell growth and the biological consequences of abnormal activity of a specific NAD+-dependent enzyme. We expressed the NTT4 gene from Chlamydiae UWE25 in Escherichia coli BW25113, as the NTT4 protein was reported as a NAD+ transporter that can specifically transport intact NAD+ across cytoplasmic membrane2. We knocked out the nadC gene responsible for de novo biosynthesis of NAD+ and constructed the strain E. coli BW25113 (ΔnadC, NTT4). It was found that NAD+ in the culture media could significantly promote the growth of BW25113 (ΔnadC, NTT4), suggesting that the NTT4 protein was functional in E. coli (Fig A, B). We then disrupted the other two genes, nadD and nadE, and obtained the strains E. coli BW25113 (ΔnadD, NTT4) and BW25113 (ΔnadE, NTT4). Cell growth of these two strains are depending on exogenous NAD+ supplemented in the media, suggesting that we have successfully engineered E. coli to hold an NAD+ auxotrophic phenotype. We are carrying out a number of experiments using these NAD+ auxotrophic strains to address some interesting questions which may not be able to do otherwise. Results will be discussed during the conference.
Language英语
Document Type会议论文
Identifierhttp://cas-ir.dicp.ac.cn/handle/321008/116038
Collection中国科学院大连化学物理研究所
Corresponding AuthorZhao ZB(赵宗保)
Recommended Citation
GB/T 7714
Zhou YJ,Hou SH,Zhang SF,et al. Construction of E. coli NAD+ auxotrophic strains and the biotechnological application thereof[C]. 待补充:待补充,2011:26-0.
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