Insights into the Hierarchical Assembly of a Chemically Diverse Peptide Hydrogel Derived from Human Semenogelin I

A peptide corresponding to a 13-residue segment of thehuman protein semenogelin I has been shown to generate a hydrogelconsisting of amyloid-like fibrils. The relative chemical diversity (comparedto synthetic de novo sequences) with 11 distinct amino acids makes thispeptide (P0) an ideal candidate for investigating the role of individualresidues in gelation. Herein, the N-terminal residues have been sequentiallyremoved to furnish a series of truncated peptides, P1−P10, ranging from 12to 3 residues in length. FTIR spectroscopy investigations reveal that P0−P6forms a β-sheet secondary structure while shorter sequences do not self-assemble. Site-specific isotope labeling of the amide backbone of P0−P2with the IR-sensitive vibrational probe 13C�O yields FTIR spectraindicative of the initial formation of a kinetic product that slowlytransforms into a structurally different thermodynamic product. The effectsof the isotopic labels on the IR spectra facilitate the assignment of parallel and antiparallel structures, which are sometimescoexistent. Additional IR studies of three PheCN-labeled P0 sequences are consistent with an H-bonded β-sheet amide core,spanning the 7 central residues. The macromolecular assembly of peptides that form β-sheets was assessed by cryo-TEM,SAXS/WAXS, and rheology. Cryo-TEM images of peptides P1−P6 display μm-long nanofibrils. Peptides P0−P3 generatehomogeneous hydrogels composed of colloidally stable nanofibrils, and P4−P6 undergo phase separation due to theaccumulation of attractive interfibrillar interactions. Three amino acid residues, Ser39, Phe40, and Gln43, were identified to beof particular interest in the truncated peptide series as the removal of any one of them, as the sequence shortens, leads to amajor change in material properties.