FIG. 4.

In vitro interaction between TYLCV CP (TYCP) and Hamiltonella GroEL (HGEL) in a pulldown assay. Polyhistidine and nonpolyhistidine in vitro-synthesized Hamiltonella (His-HGEL and HGEL, respectively), Portiera (His-PGEL and PGEL, respectively), Rickettsia (His-RGEL and RGEL, respectively), and TYLCV CP (His-TYCP and TYCP, respectively) were subjected to protein pulldown assays as described in Materials and Methods. Each pulldown assay was performed by binding one polyhistidine protein to a cobalt agarose column and immobilizing a second nonpolyhistidine protein to the column. After elution, proteins were subjected to an SDS-PAGE and Western blot analysis for detecting the proteins with anti-biotin streptavidin-HRP antibody. Hamiltonella GroEL interacted with TYLCV CP (lanes 1 and 2), but not with Rickettsia and Portiera GroELs (RGEL and PGEL, respectively) (lanes 3 to 6). Lanes 7 to 10 contain eluted 6×His-labeled recombinant proteins for size confirmation. The positions of molecular size markers (in kilodaltons) are shown to the left of the gel.