Stabilization of phosphorylated Bacillus subtilis DegU by DegR

Abstract

The production of Bacillus subtilis extracellular proteases is under positive and negative regulation. The functional role of degR, one of the positive regulators, was studied in relation to the degS and degU gene products, which belong to the bacterial two-component regulatory system. Studies with a translational fusion between the Escherichia coli lacZ and the Bacillus subtilis subtilisin (aprE) genes indicated that the stimulatory site of DegR lay upstream of position -140, with the region upstream of position -200 being the major target. It was also found that degS and degU were epistatic to degR. These results suggested some relationship among the degR, degS, and degU gene products. The DegR protein was purified to homogeneity, and its in vitro effect on the phosphorylation reaction involving DegS and DegU was studied. For this purpose, a soluble-extract system in which the formation and dephosphorylation of DegU-phosphate could be examined was devised. The addition of DegR to the soluble-extract system enhanced the formation of DegU-phosphate. The enhancing effect was found to be due to the protection of DegU-phosphate from dephosphorylation. From these results, it was concluded that the positive effect of DegR on the production of the extracellular proteases is brought about by the stabilization of DegU-phosphate, which in turn may result in the stimulation of transcription of the exoprotease genes.