The isolated catalytic domain of NIFA, a bacterial enhancer-binding protein, activates transcription in vitro: activation is inhibited by NIFL.
The NIFA protein of Klebsiella pneumoniae is required for transcription of all nif (nitrogen fixation) operons except the regulatory nifLA operon itself. NIFA activates transcription of nif operons by the alternative holoenzyme form of RNA polymerase, sigma 54-holoenzyme, in a nucleoside triphosphate (NTP)-dependent manner. NIFL antagonizes the action of NIFA in the presence of molecular oxygen or combined nitrogen. The NIFA protein of K. pneumoniae is composed of three domains: an N-terminal domain with unclear function, a central catalytic domain, and a C-terminal DNA-binding domain. We report that the isolated central domain of NIFA activates transcription in vitro and that this activation requires NTP with a hydrolyzable beta-gamma bond, as does activation by intact NIFA. Transcriptional activation by the isolated central domain has the heat lability characteristic of intact NIFA and is inhibited by NIFL. The central domain has an NTPase activity that is also heat-labile but is not inhibited by NIFL. Taken together, these results imply that NIFL interferes with contact between NIFA and sigma 54-holoenzyme.
Berger
D K
DK
Department of Plant Biology, University of California, Berkeley 94720.
Narberhaus
F
F
Kustu
S
S
D000251:Adenosine Triphosphatases / Q000235:genetics; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006358:Hot Temperature; D007711:Klebsiella pneumoniae / Q000235:genetics*; D008969:Molecular Sequence Data; D009586:Nitrogen Fixation / Q000235:genetics*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010446:Peptide Fragments; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000502:physiology*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C061338:NifA protein, Bacteria; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D012333:RNA, Messenger; D014157:Transcription Factors; C084296:nifL protein, Bacteria; D000251:Adenosine Triphosphatases