Skip to main content
Fig. 4 | Parasites & Vectors

Fig. 4

From: TBP and SNAP50 transcription factors bind specifically to the Pr77 promoter sequence from trypanosomatid non-LTR retrotransposons

Fig. 4

Binding analyses and binding kinetics of TBP and SNAP50 to the Pr77 sequence by EMSA. Increasing amounts (0.32–6.35 µM) of a (top panel) rTBP and b rSNAP50 proteins were incubated with 0.33 nM 32P-radiolabelled double-stranded Pr77 sequence (32P-dsPr77) as a probe. a Bottom panel: binding kinetics of rTBP to 32P-dsPr77. Binding data from three independent experiments, as shown in Fig. 4a and Figure S1, were fitted to the Hill model, and thermodynamic parameters Bmax (maximum percentage of bound DNA), Kd (dissociation constant) and αH (Hill coefficient) were calculated. c 32P-dsPr77 (0.33 nM) was incubated with 6.35 μM TBP, SNAP50, BSA, KMP11 and T-GP63 proteins (+), and binding reactions (+) were subjected to UV-crosslinking (XL). d 32P-dsPr77 (0.33 nM) was incubated with 6.35 μM rSNAP50 protein, and binding reactions were subjected (+) to UV-crosslinking (XL). Reactions performed with the same amount of P32-dsPr77 and without protein were subjected to UV-crosslinking (+) as a control. After incubation at 37 °C for 30 min, reactions were resolved on 6% native polyacrylamide gels. The shifted complexes formed are indicated with a black arrowhead and free 32P-dsPr77 probe with an asterisk

Back to article page