Genetic analysis on the NifW by utilizing the yeast two-hybrid system revealed that the NifW of Azotobacter vinelandii interacts with the NifZ to form higher-order complexes.
Nitrogenase is a complex metalloenzyme composed of two separately purified proteins designated the Fe-protein and the MoFe-protein. Apart from these two proteins, a number of accessory proteins are essential for the maturation and assembly of nitrogenase. Even though experimental evidence suggests that these accessory proteins are required for nitrogenase activity, the exact roles played by many of these proteins in the functions of nitrogenase are unclear. Our studies were directed to understand the role of two nif accessory proteins, the NifW and the NifZ in the biological nitrogen fixation. To accomplish this, we have utilized a genetic method, the Yeast based Two-Hybrid protein-protein interaction assay. This analysis showed that the NifW could interact with itself to make a multimeric complex. In contrast, the NifZ could not interact with itself. However, the NifZ could interact with the NifW. Previously it was shown that mutating either the NifW or the NifZ have similar effects on the activity of nitrogenase. This observation indicated that both these proteins may exert their regulation on the nitrogenase by a common pathway. Furthermore, it was suggested that the NifW plays a role in the oxygen-protection of the MoFe-protein by direct physical interaction. Our observation that the NifW can interact with itself as well as with the NifZ, suggests that the NifW and the NifZ may form a higher order complex and such a complex may be needed to exert the effects of the NifW or the NifZ on the nitrogenase activity.
Lee
S H
SH
Department of Biological Sciences, Bowling Green State University, Ohio 43403, USA.
Pulakat
L
L
Parker
K C
KC
Gavini
N
N
D000595:Amino Acid Sequence; D019677:Artificial Gene Fusion; D016948:Azotobacter vinelandii / Q000201:enzymology* / Q000235:genetics*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D005798:Genes, Bacterial; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009586:Nitrogen Fixation; D009591:Nitrogenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D001616:beta-Galactosidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D017931:DNA Primers; D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; C068360:nitrogenase stabilizing-protective protein, Bacteria; D009591:Nitrogenase; D001616:beta-Galactosidase