FeMo cofactor biosynthesis in a nifE- mutant of Rhodobacter capsulatus

Area of Science:

• Microbiology
• Biochemistry
• Molecular Biology

Background:

• The FeMo cofactor (FeMoco) is essential for nitrogen fixation in diazotrophic microorganisms.
• Mutations in nifE, a gene involved in FeMoco biosynthesis, typically result in inactive dinitrogenase.
• The specific role of NifNE proteins in FeMoco biosynthesis requires further investigation.

Purpose of the Study:

• To investigate the function of the nifE gene in the phototrophic bacterium Rhodobacter capsulatus.
• To determine if nifE mutations affect nitrogen fixation capabilities in R. capsulatus.
• To re-evaluate the role of NifNE in the FeMo cofactor biosynthetic pathway.

Main Methods:

• Generating nifE mutant strains of Rhodobacter capsulatus.
• Assessing diazotrophic growth and acetylene reduction activity in mutant strains.
• Analyzing MoFe protein expression, FeMoco incorporation, and cofactor content using EPR spectroscopy and ICP-MS.

Main Results:

• R. capsulatus nifE mutants exhibit reduced but significant acetylene reduction and diazotrophic growth.
• MoFe protein levels are decreased in nifE mutants, correlating with lower activity.
• FeMoco is incorporated into the MoFe protein in nifE mutants, albeit at lower levels than wild-type.

Conclusions:

• NifNE is not essential for FeMoco biosynthesis but contributes to higher yields of active MoFe protein.
• The findings necessitate a reconsideration of the NifNE proteins' role in the FeMoco biosynthetic pathway.
• This study demonstrates FeMoco incorporation in the absence of functional NifNE in R. capsulatus.