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. 2020 Jan 17;10:3063. doi: 10.3389/fmicb.2019.03063

FIGURE 6.

FIGURE 6

Specific binding of BrpT and BrpS to PcabA. (A,B) A 337-bp DNA fragment of the PcabA regulatory region (2.5 nM) was radioactively labeled and incubated with increasing amounts of BrpT (A) or BrpS (B) as indicated in the presence of c-di-GMP (50 μM). For competition analysis, the same but unlabeled 337-bp DNA fragment was used as a self-competitor DNA. Various amounts of self-competitor DNA were added as indicated to a reaction mixture containing the probe DNA before the addition of BrpT (A) or BrpS (B). B, bound DNA; F, free DNA. (C,D) A 337-bp DNA fragment of same PcabA regulatory region as above was labeled with 6-FAM and then used as a probe DNA. The 6-FAM-labeled probe DNA (48 nM) was incubated with increasing amounts of BrpT (C) or BrpS (D) as indicated in the presence of c-di-GMP (50 μM), and then digested with DNase I. The regions protected from DNase I cleavage by BrpT or BrpS are indicated by a white (C, BRPTB) or gray box (D, BRPSB), respectively. Nucleotide numbers shown are relative to the transcription start site of PcabA.