Fig. 4. BTI directly binds to BrpT and attenuates transcription in vitro.

(A) Microscale thermophoresis analysis demonstrating the direct interaction between BTI and BrpT. The normalized thermophoresis values of BrpT (F_norm) at varying BTI concentrations are shown. (B) In vitro transcription assay determining the effect of BTI on BrpT-mediated transcriptional activation of the brpN promoter. A template DNA containing P_brpN was transcribed in vitro with 8 μM BrpT in the presence or absence of 1 mM BTI as indicated. Relative levels of the transcripts were determined using the heights of the transcript peaks measured in arbitrary fluorescent units, shown with that of BrpT without BTI set as 1. Error bars represent the SEM from three independent experiments. Statistical significance was determined by Student’s t-test (***, p < 0.0005; *, p < 0.05).