FIG. 7.

Coimmunoprecipitation of YopN and YscB following cross-linking with DSP. Soluble extracts from the parent strain, Y. pestis KIM5-3001, grown in the presence (+) or absence (−) of calcium, the yscB deletion strain KIM5-3001.P1 (ΔyscB) grown in the absence of calcium, and the yopN deletion strain KIM5-3001.6 (ΔyopN) grown in the absence of calcium were cross-linked with DSP (1 mM) for 20 min at RT. Samples were processed as described in Materials and Methods, immunoprecipitated with antisera specific for YopN or YscB, and analyzed by SDS-PAGE and immunoblotting with antisera specific for YopN or YscB. DSP cross-links were broken prior to SDS-PAGE analysis by boiling samples in the presence of 5% β-mercaptoethanol. The locations of YopN and YscB are shown by arrows. MW, molecular size standards (45, 32, 25, 16, and 6.5 kDa). Ippt., immunoprecipitation.