title: Evidence for common sites of contact between the antisigma factor SpoIIAB and its partners SpoIIAA and the developmental transcription factor sigmaF in Bacillus subtilis.
abstract: The activity of the developmental transcription factor sigmaF in Bacillus subtilis is governed by a switch involving the dual function protein SpoIIAB. SpoIIAB is an antisigma factor that forms complexes with sigmaF and with an alternative partner protein SpoIIAA. SpoIIAB is also a protein kinase that can inactivate SpoIIAA by phosphorylating it on a serine residue. We sought to identify amino acids in SpoIIAB that are involved in the formation of the SpoIIAB-SpoIIAA complex by screening for mutants that were defective in the activation of sigmaF. This genetic screen, in combination with biochemical analysis and the construction of loss-of-side-chain (alanine substitution) mutants, led to the identification of amino acid side-chains in the N-terminal region of SpoIIAB that could contact SpoIIAA. Unexpectedly, the same amino acid side-chains (R20 and N50) that appear to touch SpoIIAA are required for binding to, and may represent sites of contact with, sigmaF. We propose that the N-terminal region of SpoIIAB forms a binding surface that is responsible for the formation of both the SpoIIAB-SpoIIAA and the SpoIIAB-sigmaF complexes, and that in some cases the same amino acid side-chains contact both partner proteins. N50 is also the defining residue of a region of amino acid sequence homology known as the N-box that is shared by SpoIIAB and related serine protein kinases, as well as by members of a mechanistically dissimilar family of protein kinases that undergo autophosphorylation at a histidine residue. We discuss the implications of this finding for the mechanism of histidine autophosphorylation.
authors.affiliation: The Biological Laboratories, Harvard University, 16 Diversity Avenue, Cambridge, MA, 02138, USA.
authors.forename: D A
authors.initials: DA
authors.lastname: Garsin
authors.forename: D M
authors.initials: DM
authors.lastname: Paskowitz
authors.forename: L
authors.initials: L
authors.lastname: Duncan
authors.forename: R
authors.initials: R
authors.lastname: Losick
mesh_terms: D000409:Alanine / Q000737:chemistry; D019943:Amino Acid Substitution; D001412:Bacillus subtilis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D016297:Mutagenesis, Site-Directed; D012808:Sigma Factor / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; C064971:FliA protein, Bacteria; D012808:Sigma Factor; D014157:Transcription Factors; C092468:spoIIR protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D000409:Alanine