Figure 4.
PreSSU Forms a High Molecular Weight Complex in Wheat Germ Lysate.
(A) A translation assay (50 μL volume) was fractionated by size exclusion chromatography calibrated with protein standards of different molecular mass (open circles). Thirty fractions of 500 μL were collected and precipitated with trichloroacetic acid, separated by SDS-PAGE, blotted, and analyzed by immunostaining with antisera raised to Hsp70 and to the 14-3-3 protein as well as by autoradiography (35S-preSSU). Radioactivity (filled diamonds) was quantified by liquid scintillation counting after dissolving the nitrocellulose in dimethyl sulfoxide.
(B) Fractions 11 to 13 and 24 to 26 were combined, and the two pools were split into equal aliquots. Combined fractions were subjected to coimmunoprecipitation by antisera to 14-3-3 (α-14-3-3) and Hsp70 (α-Hsp70) as indicated. Ten percent of each combined fraction (F) was used as internal standard. An autoradiogram is shown. PIS, control experiment using preimmune serum.