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Structural Biology Programme

Macromolecular Complexes in DNA Damage Response Group

Group Leader:  Óscar Llorca
Research highlights
Hsp90-dependent maturation of phosphatidylinositol3-kinase-like kinases (PIKKs)

The family of phosphatidylinositol-3-kinase-like kinases (PIKKs) comprises several proteins, including ATM, ATR, DNA-PKcs and mTOR, considered important molecular targets for cancer therapy. It has recently been appreciated that the assembly and maturation of active and functional PIKK complexes requires Hsp90 and a dedicated co-chaperone, the R2TP complex. Interestingly, the R2TP and Hsp90-mediated maturation of PIKKs is a regulated process in the cell, opening up an opportunity to inhibit PIKKs by interfering with their maturation. With that final goal in mind, we have started to address the structural and mechanistic understanding of human R2TP. The comparison between human and yeast R2TP complexes offers some relevant clues about the function of this complex, and in 2017, we reported the structural architecture of R2TP from yeast (Rivera-Calzada et al., Structure 2017). This work was performed in collaboration with Laurence H. Pearl ( Genome Damage and Stability Centre, University of Sussex, UK).

R2TP is one of the most complex of the various Hsp90 co-chaperones identified so far: being a multi-protein complex itself, and possessing components (RUVBL1/Rvb1 and RUVBL2/ Rvb2 in humans/yeast) that like Hsp90 possess inherent ATPase activity of their own. We have described the architecture and catalytic properties of the yeast R2TP complex using a combination of cryo-electron microscopy, structural mass spectrometry, and biochemistry. Our data provide structural and mechanistic insights of how R2TP couples an Hsp90 dimer to a PIKK kinase (FIGURE). The structural and functional characterisation of the human R2TP complex is now underway, revealing an unsuspected structural and functional complexity compared to the simpler yeast model.

Electron microscopy as a tool to help understanding human diseases as well as contributing to the development of new potential therapies

As part of our collaboration with the Santiago Rodriguez de Córdoba’s group at the Centro de Investigaciones Biológicas (CIB), we used electron microscopy (EM) methods to contribute to the understanding of how several mutations identified in patients can cause diseases linked to the complement system, a component of innate immunity. For this, we analysed the structure of components of the complement system using EM. In addition, electron microscopy was used as part of an extensive characterisation of several monoclonal antibodies with potential therapeutic and diagnostic applications in diseases related to mutations and polymorphisms in proteins of the complement system (Subias Hidalgo et al., Eur J Immunol 2017). The EM structures of each antibody bound to its target protein allowed the identification of the epitopes in the 3D structure of the target. Also, these structures provided models to explain the functional properties of the antibodies as defined by Rodriguez de Córdoba’s group. These works highlight the potential of synergising methods of structural biology with genetic, functional and clinical studies, in order to understand and cure diseases.