Overproduction and characterization of the Bacillus subtilis anti-sigma factor FlgM.

FlgM is an anti-sigma factor of the flagellar-specific
sigma (s) subunit of RNA polymerase in Bacillus subtilis,
and it is responsible of the coupling of late flagellar
gene expression to the completion of the hook-basal
body structure. We have overproduced the protein in
soluble form and characterized it. FlgM forms dimers as
shown by gel exclusion chromatography and native
polyacrylamide gel electrophoresis and interacts in
vitro with the cognate sD factor. The FlgMzsD complex is
a stable heterodimer as demonstrated by gel exclusion
chromatography, chemical cross-linking, native polyacrylamide
gel electrophoresis, and isoelectric focusing.
sD belongs to the group of sigma factors able to bind to
the promoter sequence even in the absence of core RNA
polymerase. The FlgMzsD complex gave a shift in a DNA
mobility shift assay with a probe containing a sD-dependent
promoter sequence. Limited proteolysis studies
indicate the presence of two structural motifs, corresponding
to the N- and C-terminal regions, respectively.