The Azospirillum brasilense PII and PZ
proteins, encoded by the glnB and glnZ genes respectively, are intracellular transducers of
nitrogen levels with distinct functions. The PII
protein participates in nif regulation by controlling the activity of the transcriptional regulator NifA. PII is also involved in transducing the prevailing
nitrogen levels to the Fe-
protein ADP-ribosylation system. PZ regulates negatively
ammonium transport and is involved in
nitrogenase reactivation. To further investigate the role of PII and PZ in the regulation of nitrogen fixation, broad-host-range plasmids capable of over-expressing the glnB and glnZ genes under control of the
ptac promoter were constructed and introduced into A. brasilense. The
nitrogenase activity and
nitrate-dependent growth was impaired in A. brasilense cells over-expressing the PII
protein. Using immunoblot analysis we observed that the reduction of
nitrogenase activity in cells over-expressing PII was due to partial ADP-ribosylation of the Fe-
protein under derepressing conditions and a reduction in the amount of Fe-
protein. These results support the hypothesis that the unmodified PII
protein act as a signal to the DraT
enzyme to
ADP-ribosylate the Fe-
protein in response to
ammonium shock, and that it also inhibits nif gene expression. In cells over-expressing the PZ
protein the
nitrogenase reactivation after an
ammonium shock was delayed indicating that the PZ
protein is involved in regulation of DraG activity.