Biochemical and structural studies of protein complexes involved in cancer.

摘要

Three crystal structures of protein-protein complexes involved in cancer are presented in this thesis: Smad2-MH2/SARA-SBD complex in the TGFβ signal transduction pathway, and Apaf-1 CARD/caspase-9 prodomain and Smac/XIAP-BIR3 complexes in the apoptosis pathway.; From the crystal structure of Smad2-MH2/SARA-SBD complex, we show that SARA-SBD binds to Smad2-MH2 in an extended conformation mainly through hydrophobic interactions. We identify the Smad2-MH2 and SARA-SBD residues important for this interaction. Based on a comparison between the structures of Smad2-MH2 and Smad4-MH2, we propose a model for receptor-regulated Smad recognition by the type I TGFβ receptors. Possible mechanisms for tumorigenic mutations in Smad2-MH2 are discussed.; The crystal structure of Apaf-1 CARD/caspase-9 prodomain complex reveals that Apaf-1 CARD interacts with caspase-9 prodomain through two highly charged surfaces. Recognition specificity is provided by networks of intermolecular hydrogen bonds and van der Waals interactions. Mutations of important interface residues in Apaf-1 CARD or caspase-9 prodomain eliminate or reduce complex formation. One way to promote apoptosis by activating caspase-9 in Apaf-1 deficient cancer cells is suggested.; The crystal structure of a Smac/XIAP-BIR3 complex reveals that the N-terminal four residues of Smac recognize a surface groove on XIAP-BIR3, with the first residue (Ala) making the largest contribution. We propose a mutual exclusion mechanism for the relief of XIAP-BIR3's inhibition of caspase-9 by Smac. In addition, we suggest that the Drosophila proteins Reaper/Grim/Hid use a similar mode to inhibit DIAP1 based on the sequence conservation of their N terminus with that of Smac. The peptide-binding groove on XIAP-BIR3 may be a promising drug target in cancer cells and our structure lays a foundation for efforts to design such drugs.