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. 2007 Nov 26;7(2):212–222. doi: 10.1128/EC.00361-07

FIG. 3.

FIG. 3.

CrLST8 interacts with CrTOR through its kinase domain. (A) A 25-μg portion of purified GST-CrFKBP12 was bound to rapamycin and incubated with 1 mg of Chlamydomonas total extracts in the presence or absence of the reversible cross-linker DTSSP (for details, see Materials and Methods). Fusion protein complexes were immobilized on glutathione-Sepharose 4B beads and resolved by SDS-PAGE. CrTOR and CrLST8 were detected by Western blotting. (B) Pull-down assays were performed with the kinase domain of CrTOR fused to MBP (MBP-CrTORkinase) or MBP alone and total SE from Chlamydomonas cells. About 10 μg of MBP or MBP fusion protein were incubated with 4 mg of Chlamydomonas total extract. Endogenous CrLST8 bound to MBP-kin was detected by Western blotting with CrLST8 antibody.