ADUKKIPA74 · In Meso Crystallogenesis of GPCR-protein ligand complexes
FP7 — People (Marie Curie Actions)
- Duration
- 2010-08-01 → 2012-07-31
- EU contribution
- €238,783
- Participants
- 1
- Scheme
- MC-IEF
Lines connect the coordinator with its partners.
Results in brief
In Meso Crystallogenesis of GPCR-protein ligand complexes
A summary description of the project objectives The current project has two main objectives which are described below. The first objective is to facilitate a transfer of technology involving recombinant human membrane protein expression in eukaryotic cells from the researcher to the host laboratory. The second objective is for the researcher to gain knowledge and skills in the field of in meso crystallization from the host laboratory. A description of the work performed since the beginning of the project In line with the two main objectives of the project, extensive work has been done in achieving these objectives. The researcher has set up a functional tissue culture lab that is currently being used for expression and purification of recombinant human mebrane proteins. The current expertise in this area includes maintaining eukaryotic cells such as sf9 and Hi5 insect cells in culture. At present, we can maintain long-term cultures of these cell lines for recombinant protein production purposes. The researcher also implemented laboratory protocols for the generation and amplification of recombinant baculovirus' for large-scale infection and protein production. These two aspects of the technology have been robustly tested as evidenced by the fact that we have successfully expressed and purified several human membrane proteins for structure determination purposes. The candidate has also gained theoretical and practical knowledge in the field of in meso crystallization. A description of the main results achieved so far The candidate has expressed and purified several human mebrane proteins. These proteins have been subjected to extensive crystallization trials. Of these, one has currently yielded in meso crystals that diffract to 4.0 A. The expected final results and their potential impact and use (including the socio-economic impact and the wider societal implications of the project so far). It is expected that in the near future the protein crystals diffracting to 4.0 A will be further optimized to yield the structure of this therapeutically relevant protein. It is expected that from the structure, we can glean sufficient information to design novel small-molecule drugs to treat widely prevalent diseases such as Asthama and Cardiac Heart Failure.
Data: CORDIS, © European Union
Project objective
G protein-coupled receptor (GPCR) proteins are the single largest drug target, representing 30% of all marketed drugs. All GPCRs share a basic structural core of seven transmembrane alpha-helical domains. In spite of this similarity, GPCRs have evolved to be extremely versatile receptors for extracellular messengers as diverse as biogenic amines, purines and nucleic acid derivatives, lipids, peptides and proteins, odorants, pheromones, tastants, ions like calcium and protons, and even photons. Due to their significant effect on human physiology and disease, it is crucial to understand from a structural biology perspective, the molecular basis for the interaction between a GPCR and its ligand. The recent success in the structure determination of the Beta2-adrenergic GPCR and the adenosine A2a GPCR using the in meso crystallization methodolgy paves the way for further advances in structure determination of GPCR structures. As of today, there is no structural information available that helps us in understanding the molecular basis of interaction between a GPCR and its protein ligand. In this application, we propose to perform structure-function studies on two GPCRs, CXCR4 and Formyl Peptide Receptor, that are activated by the polypeptides SDF1 and f-MLF respectively. We propose to use in a synergistic fashion two novel methodologies, the BACMAM system and in meso crystallography, to express, purify and crystallize complexes between CXCR4-SDF1 and FPR-fMLF. Information obtained from the structure determination of these complexes would go a long way in helping us understand the molecular mechanisms responsible for the signal transduction properties of GPCRs that are activated by protein ligands. In addition, this information would also help us in designing and developing better drugs aimed at treating cancer metastases, AIDS and Parkinsons disease.
Original text from CORDIS.
Participants
- THE PROVOST, FELLOWS, FOUNDATION SCHOLARS & THE OTHER MEMBERS OF BOARD, OF THE COLLEGE OF THE HOLY & UNDIVIDED TRINITY OF QUEEN ELIZABETH NEAR DUBLIN · DublinCoordinatorIreland
Links
Data: CORDIS, © European Union