Objective
Here we propose a novel hybrid strategy for the structure determination of macromolecular complexes using a state of the art mass spectrometry (MS) and cryo-electron microscopy (EM) approach. This strategy allows for the accurate fitting of crystal structures into EM-maps, but also provides structural templates for the identification of macromolecular complexes within cryo-electron tomograms (CET) by pattern recognition. It promises to establish the critical link from structural analysis of isolated protein complexes to their observation in the cell. As a promising initial target for our investigations we chose the recently identified URI complex (unconventional prefoldin RBP5 interactor), a regulator in cell growth control that acts downstream of the target of rapamycin (TOR).
The structure of the URI complex is unknown, although the crystal structures of the archaeal homologues of several of its components have been solved. Therefore, it is likely to form a striking jellyfish-shaped scaffold structure consisting of prefoldins (PDFs) with having RuvB-like helicases attached to it. To generate accurate fits of crystal structures into EM-maps, additional spatial information is advantageous. A novel cross-linking strategy using a bivalent isotope tagged chemical compound, in combination with MS can provide such information. It has been established in the host institution and allows for identification of the interface of interacting proteins at primary structure level.
We plan to combine this technology with single particle cryo-EM and the fitting of crystal structures to resolve the structure of the URI complex to pseudo atomic level. In a second step, the structure of the URI complex will be used as a template to monitor interactions with the next neighbours in the signalling pathway by pattern recognition within CET. This approach will provide a structural insight into a branch of the TOR signalling pathway that has been linked to diseases like the metabolic syndrome.
Fields of science (EuroSciVoc)
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques. See: The European Science Vocabulary.
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques. See: The European Science Vocabulary.
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Keywords
Project’s keywords as indicated by the project coordinator. Not to be confused with the EuroSciVoc taxonomy (Fields of science)
Project’s keywords as indicated by the project coordinator. Not to be confused with the EuroSciVoc taxonomy (Fields of science)
Programme(s)
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Multi-annual funding programmes that define the EU’s priorities for research and innovation.
Topic(s)
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Calls for proposals are divided into topics. A topic defines a specific subject or area for which applicants can submit proposals. The description of a topic comprises its specific scope and the expected impact of the funded project.
Call for proposal
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Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.
FP6-2005-MOBILITY-5
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Funding Scheme
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Funding scheme (or “Type of Action”) inside a programme with common features. It specifies: the scope of what is funded; the reimbursement rate; specific evaluation criteria to qualify for funding; and the use of simplified forms of costs like lump sums.
Coordinator
ZURICH
Switzerland
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