| dc.description.abstract | The human guanylyl cyclase C receptor is the target for the heat-stable entero- toxin (STa) from enterotoxigenic Escherichia coli, which is responsible for more than 200 million episodes of diarrhea and 300,000 deaths per year in developing countries. The STa toxin is currently a candidate for the generation of a toxoid vaccine, and the determination of the receptor-ligand interaction would provide invaluable information for its design. In this study, we have prepared a set of three-dimensional models for the extracellular, ligand-binding domain of the hu- man GC-C receptor (GCC-ECD), based on homology with the homologous natri- uretic peptide receptors (NPRs). The modelled GCC-ECD monomer was similar to previously published models, and the models for the dimer enabled us to iden- tify residues potentially involved in the oligomerization of the receptor, as well as the receptor-ligand interaction. Those residues are located within two regions of the GCC-ECD, from Ser75 to Ser127 and from Glu175 to Arg218. Previously pub- lished studies have shown that point mutations in the first region have an effect on ligand-binding, but the second region has not been investigated at all. Two residues that had been previously proposed as the ligand-binding residues were located within the hinge region between the two sub-domains of the GCC-ECD models. Additional candidate template structures were also obtained through threading, all belonging to the Type 1 periplasmic binding fold superfamily. Fi- nally, we have taken the first step towards the setup of in vitro interaction studies by cloning the pro-sequences for the endogenous ligands of the GC-C receptor, guanylin and uroguanylin. A fragment of the GCC-ECD was also cloned, and it was successfully expressed in E. coli. Those results provide a basis for further interaction studies, both experimentally and using bioinformatics. | en_US |
