Figure 5.

Characterization of the Stl^A236T mutant (A) Testing the Stl^A236T mutant in the Stl switch system. The structural model locates the A236T mutation at the C-terminal domain shown in blue. The HTH motif is colored yellow and the N-terminal domain is colored maroon. The expression of the Stl^A236T mutant resulted in blue colonies, verifying that it is defective in DNA binding in vivo; (B) Comparative electrophoretic mobility shift assay (EMSA) with Stl^WT and Stl^A236T. Stl^A236T is less prone to complex formation with DNA. Therefore, more of the free DNA is visible at the corresponding bands than that of Stl^WT. The comparative EMSA was repeated three times with same results; (C) Densitometry results of each of the species exhibiting different electrophoretic mobilities are shown by colored bands. The discrepancy between the wild-type and mutant Stl is quite noticeable, confirming our previous in vivo results. These data suggest that the DNA binding of Stl^A236T is compromised in vitro.