Total synthesis and structural characterization of a novel protein scaffold from the snail Biomphalaria glabrata

Abstract Disulfide-rich miniproteins constitute compact and highly stable scaffolds of growing interest for molecular and structural engineering. Schistosomins are ∼80-residue proteins conserved across gastropods that form a long-standing orphan family whose structure and biological roles have remained unknown. Here, we report the total chemical synthesis and structural characterization of a schistosomin isoform from Biomphalaria glabrata, a medically relevant intermediate host of the parasite Schistosoma mansoni. Using state-of-the-art solid-phase peptide synthesis, chemoselective peptide ligation, and controlled oxidative folding, we obtained homogeneous well folded schistosomin suitable for biophysical and structural studies. High-resolution X-ray crystallography reveals a previously undescribed disulfide-rich fold defining a new class of miniprotein scaffold. Nano differential scanning fluorimetry and circular dichroism experiments demonstrate the remarkable thermal stability of this scaffold, while molecular dynamics simulations confirm the intrinsic rigidity of its disulfide-stabilized core and show that the two naturally occurring isoforms differing by a single residue exhibit nearly indistinguishable structural and dynamic properties. Finally, transcript and protein analyses across snail tissues provide the first spatial expression map of schistosomin in a medically relevant mollusk. Together, this work establishes schistosomin as a novel and robust miniprotein scaffold and provides a structural and biological framework for exploring its function and potential applications. Competing Interest Statement The authors have declared no competing interest.