Sporulation protein SpoIVFB from Bacillus subtilis enhances processing of the sigma factor precursor Pro-sigma K in the absence of other sporulation gene products.

Processing of inactive pro-sigma K to active sigma K in the mother cell compartment of sporulating Bacillus subtilis is governed by a signal transduction pathway emanating from the forespore and involving SpoIVFB in the mother cell. Coexpression of spoIVFB and sigK (encoding pro-sigma K) genes in growing B. subtilis or Escherichia coli enhanced pro-sigma K processing in the absence of other sporulation-specific gene products. The simplest explanation of these results is that SpoIVFB is a protease that processes pro-sigma K.     

Overproduction, purification, and characterization of Bacillus subtilis RNA polymerase sigma A factor.

By use of a T7 expression system, large amounts of active Bacillus subtilis RNA polymerase sigma A factor were produced in Escherichia coli cells. This overproduced protein was found in the form of inclusion bodies and constituted 40% of the total cellular protein. Because of the ease of isolation of the inclusion bodies and the acidic properties of sigma A, the protein was purified to more than 99% purity and the yield was about 90 mg/liter of culture. Gel mobility, antigenicity, specificity of promoter recognition, and N-terminal amino acid sequence of the overproduced sigma were found to be the same as those of native sigma A. Partial proteolysis analysis of sigma A protein suggested the presence of a protease-sensitive surface region in the C-terminal part of the sigma A protein. The promoter -10 binding region of sigma A was less sensitive to proteases and was probably involved in a hydrophobic, tightly folded domain of sigma A protein.