Figure 8.
Specific bindings of HlyU and CRP to PplpA. A 461-bp DNA fragment of the plpA-regulatory region was radioactively labeled and then used as a probe DNA. A and B, the radiolabeled probe DNA (5 nm) was incubated with increasing amounts of HlyU (A) and CRP (B) as indicated. For competition analysis, the same but unlabeled 461-bp DNA fragment was used as a self-competitor DNA. Various amounts of the self-competitor DNA were added to a reaction mixture containing the 5 nm labeled DNA before the addition of 40 nm HlyU (A) and 80 nm CRP (B) as indicated. B, bound DNA; F, free DNA. C and D, the same radiolabeled probe DNA (25 nm) was reacted with increasing amounts of HlyU (C) and CRP (D) as indicated. The regions protected from DNase I cleavage by HlyU and CRP are indicated by gray boxes (HLYUB1, HLYUB2, and HLYUB3) and a white box (CRPB), respectively. Lanes C, T, A, and G represent the nucleotide sequencing ladders of pKK1505. Nucleotide numbers shown are relative to the transcription start site of plpA.