Conotoxins are disulfide-rich peptides isolated from the venoms of marine cone snails. In recent times, cone snails have become one of the most significant treasure troves of lead molecules for pharmaceutical development. Many conotoxins are highly modified with post-translational modifications such as hydroxylation of proline, C-terminal amidation, and glycosylation, amongst others. These modifications can alter the peptide's charge, size, and hydrophobicity, thereby influencing its interaction with target ion channels and modulating its potency and selectivity. In addition, PTMs can also affect the conformational stability and folding of the peptide, which can further impact its biological activity. Given the limited knowledge of the structural implications of post-translational modifications (PTMs) with emphasis on proline hydroxylation and C-terminal amidation, the specific effect of PTMs on the structure and function of conotoxins needs to be investigated.

We reasoned to expand the current knowledge by elucidating the structural and functional roles of PTMs in the Mu (µ) pharmacological family of conotoxin. Consequently, we synthesized peptides isolated from marine cone snails of the Mu (µ) pharmacological family.

These conotoxins were selected because they contain naturally occurring PTMs in the peptide sequence, with modifications in their proline residue. While the structures and the pharmacology of the native variants of these conotoxins have been described, there is no information about how the PTMs contribute to their features. Hence our objective was to synthesize a series of TIIIA and PIIIA peptides bearing PTM in the form of proline hydroxylation, C-terminal amidation, and their unmodified variants with the intent of ascertaining the role of proline hydroxylation and C-terminal amidation in structural and functional activity. Consequently, this study provides novel insights and better clarity on whether the modification interacts directly with the binding site, changes the conotoxin structure, or affects the folding kinetics and biological activities of the conotoxins.