FIG. 8.
Effect of heparin on PKR dimerization. Flag-tagged and nontagged wt and L75E PKR proteins were synthesized by cotranslation in the reticulocyte lysate; 5 μl of in vitro-translated, 35S-labeled proteins were immunoprecipitated with Flag MAb-agarose either in the absence (lanes 3 to 5, 11, and 12) or in the presence (lanes 6 to 8, 13, and 14) of heparin (10 U/ml). The immunoprecipitated proteins were analyzed by SDS-PAGE on an 8% gel followed by fluorography. Lanes 1, 3, and 6, nontagged wt PKR alone; lanes 2, 4, and 7, cotranslated nontagged wt PKR and Flag-tagged wt PKR; lanes 5 and 8, Flag-tagged wt PKR alone; lanes 9, 11, and 13, nontagged L75E PKR alone; lanes 10, 12, and 14, cotranslated nontagged L75E PKR and Flag-tagged L75E PKR. Lanes 1, 2, 9, and 10 represent total proteins; lanes 3 to 8 and 11 to 14 represent immunoprecipitated (IP) proteins. Positions of nontagged and Flag-tagged proteins are as indicated.