FIG. 4.

Phosphorelay dephosphorylation assay. Purified Spo0E-like proteins of B. anthracis were tested in vitro for their ability to dephosphorylate the B. subtilis phosphorelay. KinA (0.2 μM), Spo0F (2.5 μM), Spo0B (0.2 μM), and Spo0A (2.5 μM) were incubated in the presence of [γ-³²P]ATP for 30 min. The reaction mixture was then divided in two aliquots: one was incubated with the corresponding Spo0E protein buffer (−), and the other was incubated with buffer plus the Spo0E protein (+). Aliquots were withdrawn at the indicated times. BA1877 (A), BA0046 (B), BAYisI (C), and BA2416 (D) proteins purified from E. coli were used at a 5 μM final concentration. The position of the phosphorylated proteins is indicated by the arrows.