Figure 5.

In vitro and in vivo protein–protein interaction of DevR and PhoP RRs. A, the interaction between M. tuberculosis DevR–PhoP_Rv (black bars) and DevR–PhoP_Ra (gray bars) proteins was assessed in vitro in an ELISA format using 10 pmol of purified PhoP_Rv/PhoP_Ra proteins as described under “Experimental procedures.” The background values from the control wells were subtracted from the test well readings, and ΔA₄₅₀ values are plotted. ** and ***, p < 0.01 and 0.001, respectively, for the difference in interaction between DevR–PhoP_Rv and DevR–PhoP_Ra proteins. B, in vivo mycobacterial protein fragment complementation assay. Shown is M. smegmatis transformed with the following pairs of plasmid constructs encoding recombinant fusion proteins: recombinant PhoP_Rv-[F3]/DevR-[F1,2] (1) and PhoP_Ra-[F3]/DevR-[F1,2] (2). Positive controls (3 and 4) and negative control (5) were streaked on Kan + Hyg (left) and Kan + Hyg + TRIM plates (right). One representative picture from three independent experiments is shown. Error bars, S.D.