FIG. 2.

ChxR forms homodimers in vivo. Recombinant ChxR forms homodimers in vitro; however, the in vivo oligomeric state of ChxR was unknown. (A) To determine whether the primary amine chemical cross-linker DSS could capture ChxR homodimers, increasing concentrations (2.2, 3.5, 7, 14, and 21 μM) of purified, recombinant ChxR were incubated with 500 μM DSS. As a control, 21 μM ChxR was not incubated with DSS. Denatured samples were separated by SDS-PAGE and observed by Coomassie staining. (B) At 12, 24, and 36 hpi, C. trachomatis were enriched from infected L929 cells, and the relative amount of ChxR present was assayed by an immunoblot with polyclonal-monospecific antibodies against ChxR (αChxR). The alpha subunit of RNA polymerase (αRpoA) was used to normalize the amount of chlamydial protein (ChxR) each time point. (C) To test dimer formation in vivo, 10 mM DSS was added to uninfected host cells (Mock) or C. trachomatis-enriched lysates at 30 hpi (Infected). The samples were separated by SDS-PAGE, and an immunoblot was performed with antibodies against ChxR.