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Home    Υποκλωνοποίηση του γονιδίου του τύπου 2 υποδοχέα του CRF και φαρμακολογικός χαρακτηρισμός του υποδοχέα  

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Identifier 000403387
Title Υποκλωνοποίηση του γονιδίου του τύπου 2 υποδοχέα του CRF και φαρμακολογικός χαρακτηρισμός του υποδοχέα
Alternative Title Subcloning the type 2 CRF receptor gene and pharmacological characterization of receptor
Author Κανελλόπουλος, Παναγιώτης
Thesis advisor Λιαπάκης, Γεώργιος
Reviewer Θερμού, Κυριακή
Βενυχάκη, Μαρία
Abstract Corticotropin releasing factor (CRF) is a 41 amino acid peptide, which regulates the function of the central nervous system (CNS) and other systems as well, thus playing an important role in the maintenance of the homeostasis. CRF exerts its multiple effects through its interaction with the type 1 (CRF1R) and type 2 (CRF2R) receptors. These receptors belong to the family of G-protein coupled receptors (GPCRs). CRF receptors, as all GPCRs, consist of extracellular and intracellular regions and seven transmembrane domains (TMs). CRF1R and CRF2R have different binding affinities for various CRF analogs. In specific, CRF1R and CRF2R have similar binding affinities for sauvagine and urocortin I. In contrast, urocortins II and III bind specifically to CRF2R, whereas all synthetic non-peptide antagonists are CRF1R-selective. Interestingly, all the TM amino acids of CRF1R, which have been shown in a recent crystallographic study to bind the CRF1R-selective non-peptide antagonists, are totally conserved in the CRF2R. This leads to the hypothesis that the amino acids of CRF1R that contact non-peptide molecules have different orientations in CRF2R, such as the high affinity binding of these molecules to CRF2R is not feasible. However the absence of structural information for the CRF2R hampers the verification of this hypothesis. In addition, the lack of structural information for this receptor impedes the determination of the amino acids that contact various ligands, which is the first step for the design of new CRF2R-selective molecules. To elucidate the structure and function of CRF2R, we must mutate its amino acids to different ones and determine the pharmacological properties of mutant receptors. To accomplish this, we must first create an efficient plasmid containing the CRF2R gene. In the present study we subcloned the CRF2R gene into the pCIN4 vector, thus creating the CRF2R/pCNI4 plasmid. Next we determine the functional properties of the protein encoded by the CRF2R/pCNI4 plasmid by expressing this protein (hCRF2R) into HEK cells and pharmacologically characterizing the hCRF2R. The CRF2R/pCNI4 plasmid allows the easier, faster and more reliable expression of CRF2R and possibly its mutants that will be created in our laboratory in order to elucidate the structure and function of this receptor. Determination of the structure and function of CRF2R will lead to the design of new CRF2R-selective analogs. These analogs will be extremely useful tools for the elucidation of the role of CRF2R in the function of CNS and other systems as well and possibly for the treatment of several pathological conditions in which the CRF2R is implicated.
Language Greek
Subject Εκλυτικός παράγοντας της κορτικοτροπίνης
Issue date 2016-12-13
Collection   School/Department--School of Medicine--Department of Medicine--Post-graduate theses
  Type of Work--Post-graduate theses
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