Fig. 3.

BglG becomes phosphorylated in vitro by HPr (A) as well as by FruB (B). PEP-dependent phosphorylation assays were carried out containing purified His₁₀–EI, His₁₀–HPr, Strep–BglG, or Strep–S_tag–BglG as indicated. In the assays shown in B, His₁₀–HPr was replaced by His₁₀–FruB. Assays were carried out using either [³²P]-PEP (Upper) or 1 μM cold PEP (Lower). Proteins were subsequently separated by denaturing gel electrophoresis using 15% (A) or 12% polyacrylamide gels (B). Gels were analyzed by phosphoimaging (Upper) or staining with Coomassie brilliant blue (Lower). Positions of the molecular weight marker are given at Left.