FIG 5.

Scc4 or Scc1 directly interacted with SycN and YscB in a bacterial two-hybrid assay. (a) Schematic showing a transcription activation-based bacterial two-hybrid assay. In this system, protein-protein interactions between the indicated hybrid proteins in a reporter E. coli strain, FW102 OL2-62, are quantified by measuring β-galactosidase activities. (b) Immunoblot analysis of protein expression in strain FW102 OL2-62 carrying pBR-yscB, pBR-sycN, or pBRα (vector control). Rabbit anti-YscB or anti-SycN antibody was used to probe proteins as indicated. (c and d) Results of β-galactosidase activities illustrating the protein-protein interactions. Note that SycN-Scc4 interactions (c) were stronger than those of YscB-Scc4 (d). Protein expression in culture was induced by the addition of IPTG (50 μM). Result obtained from the negative control (CI) represents the background. Shown are the means ± SD from three biological replicates, each in technical triplicates.