The Bacillus subtilis GTP Binding Protein Obg and Regulators of the ς(B) Stress Response Transcription Factor Cofractionate with Ribosomes

Obg, an essential GTP binding protein of Bacillus subtilis, is necessary for stress activation of the ς(B) transcription factor. We investigated Obg's cellular associations by differential centrifugation of crude B. subtilis extracts, using an anti-Obg antibody as a probe to monitor Obg during the fractionation, and by fluorescent microscopy of a B. subtilis strain in which Obg was fused to green fluorescent protein. The results indicated that Obg is part of a large cytoplasmic complex. In subsequent analyses, Obg coeluted with ribosomal subunits during gel filtration of B. subtilis lysates on Sephacryl S-400 and specifically bound to ribosomal protein L13 in an affinity blot assay. Probing the gel filtration fractions with antibodies specific for ς(B) and its coexpressed regulators (Rsb proteins) revealed coincident elution of the upstream components of the ς(B) stress activation pathway (RsbR, -S, and -T) with Obg and the ribosomal subunits. The data implicate ribosome function as a possible mediator of the activity of Obg and the stress induction of ς(B).