title: Interspecies Transfer and Regulation of Pseudomonas stutzeri A1501 Nitrogen Fixation Island in Escherichia coli.
abstract: Until now, considerable effort has been made to engineer novel nitrogen-fixing organisms through the transfer of nif genes from various diazotrophs to non-nitrogen fixers; however, regulatory coupling of the heterologous nif genes with the regulatory system of the new host is still not well understood. In this work, a 49 kb nitrogen fixation island from P. stutzeri A1501 was transferred into E. coli using a novel and efficient transformation strategy, and a series of recombinant nitrogen-fixing E. coli strains were obtained. We found that the nitrogenase activity of the recombinant E. coli strain EN-01, similar to the parent strain P. stutzeri A1501, was dependent on external ammonia concentration, oxygen tension, and temperature. We further found that there existed a regulatory coupling between the E. coli general nitrogen regulatory system and the heterologous P. stutzeri nif island in the recombinant E. coli strain. We also provided evidence that the E. coli general nitrogen regulator GlnG protein was involved in the activation of the nif-specific regulator NifA via a direct interaction with the NifA promoter. To the best of our knowledge, this work plays a groundbreaking role in increasing understanding of the regulatory coupling of the heterologous nitrogen fixation system with the regulatory system of the recipient host. Furthermore, it will shed light on the structure and functional integrity of the nif island and will be useful for the construction of novel and more robust nitrogen-fixing organisms through biosynthetic engineering.
authors.affiliation: Key Laboratory of Bio-resources and Eco-environment Ministry of Education, College of Life Sciences, Sichuan University, Chengdu, Sichuan 610064, P.R. China.|Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.
authors.forename: Yunlei
authors.initials: Y
authors.lastname: Han
authors.affiliation: CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, P.R. China.
authors.forename: Na
authors.initials: N
authors.lastname: Lu
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.|CAS Key Laboratory for Biological Effects of Nanomaterials and Nanosafety, National Center for Nanoscience and Technology, Beijing 100190, P.R. China.
authors.forename: Qinghua
authors.initials: Q
authors.lastname: Chen
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.
authors.forename: Yuhua
authors.initials: Y
authors.lastname: Zhan
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.|Institute of Plant Protection and Microbiology, Zhejiang Academy of Agricultural Sciences, Zhejiang 310021, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Lu
authors.affiliation: CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, P.R. China.
authors.forename: Baoli
authors.initials: B
authors.lastname: Zhu
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.
authors.forename: Min
authors.initials: M
authors.lastname: Lin
authors.affiliation: Key Laboratory of Bio-resources and Eco-environment Ministry of Education, College of Life Sciences, Sichuan University, Chengdu, Sichuan 610064, P.R. China.
authors.forename: Zhirong
authors.initials: Z
authors.lastname: Yang
authors.affiliation: Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, P.R. China.
authors.forename: Yongliang
authors.initials: Y
authors.lastname: Yan
mesh_terms: D000641:Ammonia / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D053263:Gene Regulatory Networks; D022761:Gene Transfer, Horizontal; D044404:Genomic Islands; D053858:Metabolic Networks and Pathways / Q000235:genetics*; D009586:Nitrogen Fixation; D010100:Oxygen / Q000378:metabolism; D044223:Pseudomonas stutzeri / Q000235:genetics* / Q000378:metabolism; D013696:Temperature
chemical_list: D000641:Ammonia; D010100:Oxygen
keywords: Escherichia coli; Pseudomonas stutzeri A1501; biological nitrogen fixation; horizontal gene transfer; nitrogen fixation island