FIG. 5.

Ansamycin benzoquinones block kinase activity of Gcn2 in vitro. Gcn2 was translated at 30°C in rabbit reticulocyte lysate. Macbecin I (Mc) and GA were added during synthesis, where indicated. After immunoprecipitation with an anti-Gcn2 antiserum, the samples were used for in vitro kinase assays and electrophoresed on an SDS-polyacrylamide gel electrophoresis gel. The two panels represent the same gel with differential detection of incorporated ³⁵S (upper panel) and ³²P (lower panel) by using two sheets of X-ray film. The numbers above the panels represent the concentrations of compounds used (in micromolar). Note that the slightly reduced levels of immunoprecipitated Gcn2 in the presence of Mc were specific to this particular experiment and are not generally observed. −, no drug added.