Expression of nitrogen fixation genes in foreign hosts. Assembly of nitrogenase Fe protein in Escherichia coli and in yeast.
In Klebsiella pneumoniae, the nifH gene encodes the Fe protein (Kp2) polypeptide that is assembled into a homodimer responsible for the reduction of nitrogenase. Escherichia coli or the yeast Saccharomyces cerevisiae, transformed with the K. pneumoniae nifH gene in suitable expression vectors, synthesize the Fe protein polypeptide. This study examines the assembly of the nifH gene product into its characteristic dimeric structure in E. coli and in yeast. Immunoblotting methods, as well as 55Fe2- labeling of K. pneumoniae were employed to detect native nitrogenase components in cell lysates. E. coli and yeast transformants contained a protein similar to native Kp2 in its immunoreactivity, apparent molecular weight, and lability in the presence of oxygen or MgATP. While in E. coli the co-introduction of nifH and nifM resulted in enhanced levels of the nifH product, it appears that the nifH gene product alone is sufficient for the assembly of an Fe protein-like structure in foreign prokaryotic and eukaryotic hosts.
D000255:Adenosine Triphosphate / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D007163:Immunosorbent Techniques; D007506:Iron-Sulfur Proteins / Q000096:biosynthesis; D007711:Klebsiella pneumoniae / Q000235:genetics*; D046911:Macromolecular Substances; D009586:Nitrogen Fixation; D009591:Nitrogenase / Q000096:biosynthesis*; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics
D007506:Iron-Sulfur Proteins; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D009591:Nitrogenase