ASSEMBLING OF THE EXTRACELLULAR CRF RECEPTOR DOMAINS BY MEANS OF CHEMICAL AND EMZYMATIC LIGATIONS

摘要

Ligand binding is the initial step of stimulation for G protein-coupled receptors (GPCRs), transmitting signals from the extracellular environment to the interior of the cell. Understanding how ligands bind to GPCRs at the molecular level is hampered by a lack of appropriate proteins for high resolution structural analysis, and the search for new ligands is limited by the need of cellular systems which exhibit appropriate receptors. Extracellular receptor domains (ECDs) play often a major role for ligand binding and can be, therefore, interesting tools to screening potential ligands and obtain first structural information on ligand binding mechanisms. Whilst single ECDs are easily available, chemically or by expression in E. coli, a construct of the four ECDs of a 7tm-GPCR has not been achieved yet. Here, we report on the first semi-synthesis of such a 23 kD-constract, consisting of ECD1-4 of the 7tm-GPCR Corticotropin-Releasing Factor receptor.