FIG. 2.

DdAtg1^K36A lacks kinase activity. Plasmids expressing GST-tagged DdAtg1 and DdAtg1^K36A in wild-type DH1 (lanes 1 and 2) or atg1-null cells (lane 3). The proteins were purified by a pull-down with glutathione beads and assayed for autophosphorylation (top). The right lanes (lanes 4 to 6) show that a deletion of the C-terminal portion of the Atg1 kinase does not affect the kinase activity. Western blots (bottom) with anti-GST antibodies of the same samples are shown. DdAtg1^K36A is not phosphorylated in the atg1-null background but is phosphorylated in the DH1 background (compare lane 3 to lane 2).