STa peptide analogs for probing guanylyl cyclase C

Guanylyl cyclase C (GCC), universally over-expressed on primary and metastatic colorectal carcinoma cells, is activated by endogenous ligands guanylin and uroguanylin and exogenous 18 residue heat-stable enterotoxins (STa) produced by diarrheagenic bacteria. Two 12 residue STa analogs with different combinations of two interlocked disulfide bonds, peptide 3 and peptide 6, were synthesized by orthogonal solid phase synthesis routes. Peptide 3 and peptide 6 bound GCC with a rank order potency of STa > peptide 3 > peptide 6. Peptide 6 behaved as an agonist. Conversely, peptide 3 was an antagonist of GCC at nM concentrations, but an agonist at μM concentrations. These observations demonstrate the utility of this novel solid phase synthesis route for generating peptides with complex disulfide bonds. Furthermore, the results reveal that the minimum toxic domain of ST encompasses 12 amino acid residues. Moreover, they identify a specific sequence of this diarrheagenic toxin that serves at low concentration as an antagonist for GCC.