Figure 3

rrd1 Δ mutants are unable to induce conformational changes to the GST-CTD in response to rapamycin. A) Silver stained gel of purified GST and GST-CTD. The indicated strains carrying either the GST (lanes 1 and 2) or GST-CTD expressing plasmid (lanes 3-6) were untreated (-) or treated (+) with rapamycin (RAP) (200 ng/ml for 30 min). B) Far-UV circular dichroism (CD) spectral analysis of purified GST-CTD. The purified GST-CTD (0.45 μM) was derived from the parent strain (triangle) or rrd1 Δ mutant (circle) that were untreated (opened symbol) or treated (closed symbol) with rapamycin. C) Far-UV CD spectral analysis of purified GST-CTD. The purified GST-CTD (0.45 μM) was derived from the rrd1 Δ mutant carrying the empty vector (circle) or the pRRD1 plasmid (triangle) that were untreated (opened symbol) or treated (closed symbol) with rapamycin. D) CD analysis of purified GST (0.76 μM) derived from untreated (opened symbol) and rapamycin treated (closed symbol) parent cells as above. Results shown are the averages of two independent experiments. E) Limited proteolysis of purified GST-CTD derived from parent cells untreated or treated with rapamycin. The purified GST-CTD was subjected to partial chymotrypsin digestion and analyzed by silver staining. Results shown are representative of two independent experiments.