中国科学院大连化学物理研究所机构知识库
Advanced  
DICP OpenIR  > 中国科学院大连化学物理研究所  > 期刊论文
题名: Determining the Extremes of the Cellular NAD(H) Level by Using an Escherichia coli NAD(+)-Auxotrophic Mutant
作者: Zhou, Yongjin1, 3;  Wang, Lei1, 3;  Yang, Fan1;  Lin, Xinping1, 3;  Zhang, Sufang1;  Zhao, Zongbao K.1, 2
刊名: APPLIED AND ENVIRONMENTAL MICROBIOLOGY
发表日期: 2011-09-01
DOI: 10.1128/AEM.00630-11
卷: 77, 期:17, 页:6133-6140
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology ;  Life Sciences & Biomedicine
类目[WOS]: Biotechnology & Applied Microbiology ;  Microbiology
研究领域[WOS]: Biotechnology & Applied Microbiology ;  Microbiology
英文摘要: NAD (NAD(+)) and its reduced form (NADH) are omnipresent cofactors in biological systems. However, it is difficult to determine the extremes of the cellular NAD(H) level in live cells because the NAD(+) level is tightly controlled by a biosynthesis regulation mechanism. Here, we developed a strategy to determine the extreme NAD(H) levels in Escherichia coli cells that were genetically engineered to be NAD(+) auxotrophic. First, we expressed the ntt4 gene encoding the NAD(H) transporter in the E. coli mutant YJE001, which had a deletion of the nadC gene responsible for NAD(+) de novo biosynthesis, and we showed NTT4 conferred on the mutant strain better growth in the presence of exogenous NAD(+). We then constructed the NAD(+)-auxotrophic mutant YJE003 by disrupting the essential gene nadE, which is responsible for the last step of NAD(+) biosynthesis in cells harboring the ntt4 gene. The minimal NAD(+) level was determined in M9 medium in proliferating YJE003 cells that were preloaded with NAD(+), while the maximal NAD(H) level was determined by exposing the cells to high concentrations of exogenous NAD(H). Compared with supplementation of NADH, cells grew faster and had a higher intracellular NAD(H) level when NAD(+) was fed. The intracellular NAD(H) level increased with the increase of exogenous NAD(+) concentration, until it reached a plateau. Thus, a minimal NAD(H) level of 0.039 mM and a maximum of 8.49 mM were determined, which were 0.044 x and 9.6 x those of wild-type cells, respectively. Finally, the potential application of this strategy in biotechnology is briefly discussed.
关键词[WOS]: SACCHAROMYCES-CEREVISIAE ;  NADH AVAILABILITY ;  SEQUENCE-ANALYSIS ;  GLYCOLYTIC FLUX ;  NAD(P)(H) POOL ;  METABOLISM ;  ENHANCEMENT ;  TRANSPORTER ;  EXPRESSION ;  MEMBRANE
语种: 英语
WOS记录号: WOS:000294205700036
Citation statistics: 
内容类型: 期刊论文
URI标识: http://cas-ir.dicp.ac.cn/handle/321008/142672
Appears in Collections:中国科学院大连化学物理研究所_期刊论文

Files in This Item:

There are no files associated with this item.


作者单位: 1.Chinese Acad Sci, Dalian Inst Chem Phys, Dalian 116023, Peoples R China
2.Dalian Natl Lab Clean Energy, Dalian 116023, Peoples R China
3.Chinese Acad Sci, Grad Univ, Beijing 100049, Peoples R China

Recommended Citation:
Zhou, Yongjin,Wang, Lei,Yang, Fan,et al. Determining the Extremes of the Cellular NAD(H) Level by Using an Escherichia coli NAD(+)-Auxotrophic Mutant[J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY,2011,77(17):6133-6140.
Service
 Recommend this item
 Sava as my favorate item
 Show this item's statistics
 Export Endnote File
Google Scholar
 Similar articles in Google Scholar
 [Zhou, Yongjin]'s Articles
 [Wang, Lei]'s Articles
 [Yang, Fan]'s Articles
CSDL cross search
 Similar articles in CSDL Cross Search
 [Zhou, Yongjin]‘s Articles
 [Wang, Lei]‘s Articles
 [Yang, Fan]‘s Articles
Related Copyright Policies
Null
Social Bookmarking
  Add to CiteULike  Add to Connotea  Add to Del.icio.us  Add to Digg  Add to Reddit 
所有评论 (0)
暂无评论
 
评注功能仅针对注册用户开放,请您登录
您对该条目有什么异议,请填写以下表单,管理员会尽快联系您。
内 容:
Email:  *
单位:
验证码:   刷新
您在IR的使用过程中有什么好的想法或者建议可以反馈给我们。
标 题:
 *
内 容:
Email:  *
验证码:   刷新

Items in IR are protected by copyright, with all rights reserved, unless otherwise indicated.

 

 

Valid XHTML 1.0!
Powered by CSpace