Establishing differential gene expression in sporulating Bacillus subtilis: phosphorylation of SpoIIAA (anti-anti-sigmaF) alters its conformation and prevents formation of a SpoIIAA/SpoIIAB/ADP complex.
Sigma-factor F (sigmaF) is a key transcription factor that initiates prespore development in Bacillus subtilis. Its activity is controlled by an anti-sigma factor, SpoIIAB, which is also a protein kinase that phosphorylates the anti-anti-sigma factor SpoIIAA. We have examined our earlier prediction that SpoIIAA must undergo a major change in its properties when phosphorylated. Upon gel filtration in the presence of ADP, SpoIIAA-P was eluted from a Superdex column much later than SpoIIAB, whereas SpoIIAA was coeluted with SpoIIAB, indicating the formation of a protein/protein complex. The complex contained ADP, and had two monomers of SpoIIAA to each SpoIIAB dimer. Its dissociation constant was 13 mu M. Gel permeation on high-performance liquid chromatography (HPLC) suggested an apparent molecular mass for SpoIIAA-P which was much higher (23.5 kDa) than that of SpoIIAA (15.8 kDa), but Ferguson plots showed that SpoIIAA-P was not a phosphorylated dimer of SpoIIAA. Our tentative conclusion, that SpoIIAA and SpoIIAA-P differ markedly in conformation, was confirmed by the results of partial digestion with chymotrypsin.
Magnin
T
T
Microbiology Unit, Department of Biochemistry, University of Oxford, UK.
Lord
M
M
Errington
J
J
Yudkin
M D
MD
D000244:Adenosine Diphosphate / Q000378:metabolism*; D001412:Bacillus subtilis / Q000502:physiology*; D001426:Bacterial Proteins / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008957:Models, Genetic; D010766:Phosphorylation; D011487:Protein Conformation; D012808:Sigma Factor; D013171:Spores, Bacterial; D014157:Transcription Factors
D001426:Bacterial Proteins; C064971:FliA protein, Bacteria; D012808:Sigma Factor; D014157:Transcription Factors; C092468:spoIIR protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D000244:Adenosine Diphosphate