| Products/Services Used | Details | Operation |
|---|---|---|
| Codon Optimization | The genes encoding the A. vinelandii NifH (with or without an N-terminal polyhistidine tag), NifM, NifE (with an N-terminal polyhistidine tag), and NifN proteins and the M. acetivorans NifS3, NifU3, and NifB proteins were codon optimized for E. coli expression, synthesized, and cloned into pCDFDuet-1 (carrying the Ma_nifS3,U3,B and Av_nifE,N genes), pRSFDuet-1 (carrying the Av_nifH,M genes), respectively (GenScript, Piscataway, NJ). Subsequently, these constructs were transformed into E. coli strain MY21, which was derived from E. coli strain BL21(DE3) but contained a deletion of iscR, the gene encoding the FeS cluster regulator, in the genome. | Get A Quote |
The heterologous synthesis of a nitrogen-fixing system in a non-diazotrophic organism is a long-sought-after goal because of the crucial importance of nitrogenase for agronomy, energy, and the environment. Here, we report the heterologous synthesis of a two-component nitrogenase analog from Azotobacter vinelandii, which consists of the reductase component (NifH) and the cofactor maturase (NifEN), in Escherichia coli. Metal, electron paramagnetic resonance, and activity analyses verify the cluster composition and functional competence of the heterologously expressed NifH and NifEN. Nuclear magnetic resonance, nanoscale secondary ion mass spectrometry, and growth experiments further illustrate the ability of the ... More
