Evidence indicating that the extracellular loops of the mouse MC5 receptor do not participate in ligand binding

Abstract

The mMC5 receptor was cloned from a genomic library, mutated in the extracellular loops (EL's), expressed and tested for binding to melanocyte stimulating hormone (MSH) peptides. The EL's show low amino acid homology within the MC receptor family. Two mutants of the mMC5 receptor were created in order to investigate the participation of these regions in ligand binding. The EL1 and EL3 were separately altered by multiple mutagenesis so that their amino acid sequences became identical with the hMC1 receptor. The mutants were expressed in COS cells and found to bind peptide ligands in the same fashion as the wild type mMC5 receptor clone. The results indicate that the amino acids that were mutated in the mMC5 receptor do not participate in binding of MSH peptides. Comparison of the wild type mMC5 receptor with the hMC5 receptor showed that it has the same potency order for the MSH peptides but considerably higher affinity than the hMC5 receptor.