SIGT-OR-G

Signal transduction of Olfactory Receptors to their G protein

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 Obiettivo del progetto (Objective)

'G protein coupled receptors form the largest and most important class of cell surface receptors with a seven transmembrane fold. These proteins mediate diverse cell responses as reaction to input signals like hormones, peptides, light and endogenous ligands. Due to their central role, most drugs on the market target these receptors. In 2011 and 2012 incredible advance has been published in terms of X-ray structural data on GPCRs. Nevertheless, the actual mode of action and structural rearrangements remain poorly understood. Here the field of olfaction is lagging behind even more with few examples of purified receptors and no structural data. This project proposes to characterize the interaction of an Olfactory Receptor (OR) to its G protein Gaolf from the Gas family. As basis a structural model of the OR will be evaluated by Cys scanning to identify the crucial interaction region of ILC2. In parallel the G protein will be crystalized to enable co-crystallization experiments with loop fragments. Based on these information peptides resembling ICL2 will be generated and examined for their ability to activate the G protein by fluorescent GTP analogs. The peptide will be kept in a loop-like conformation by a chemical, bivalent molecule. In a final step an azo compound will be incorporated in this molecule enabling to change the length in a light dependent manner to exert control over G protein activation by light.'

Introduzione (Teaser)

Understanding how cells respond to extracellular signals is central to biomedical research. European scientists set out to dissect the initial events of receptor-mediated signalling.

Descrizione progetto (Article)

Receptors that are coupled to G proteins (GPCRs) constitute one of the most important classes of cell surface receptors. They mediate diverse cellular processes in response to hormones, peptides or light. Given their central role, a large number of drugs in the market have been designed to target these receptors.

Although there is a great deal of structural information on GPCRs, the mechanistic details of the downstream transduction events remain elusive. Scientists on the EU-funded SIGT-OR-G (Signal transduction of olfactory receptors to their G protein) project focused on the interaction of olfactory receptors (OR) and its G proteins to address this lack of knowledge.

Considerable efforts went into the optimisation of all the important steps implicated in the functional expression and purification of ORs and the G protein. To facilitate their studies, researchers first expressed human OR5P3 in cells and then directly observed the specific binding of a fluorescent odorant. To investigate the interaction between the OR and its G protein, they synthesised a fluorescent peptide predicted to be directly involved in the OR-G complex.

SIGT-OR-G also tested another approach that involved the isolation of ORs together with the entire signalling machinery in vesicles. For this purpose, they developed a procedure for purification and characterisation of such cell-derived vesicles.

Collectively, the methods and tools developed during the SIGT-OR-G will aid the future investigation of ORs and other transmembrane receptors. In the long term the generated information could prove useful in the design of novel pharmaceuticals.