A fragment of 12S seed storage protein of Arabidopsis forms twisted cross beta-sheet rich amyloid fibrils

Abstract

Plant seed storage proteins (SSPs) serve as a nutrient source and are suggested to be crucial for seed survival during dormancy and desiccation. They form highly stable and environmentally stress-resistant amyloid struc tures. SSP amyloids are gaining significant attention for fabricating sustainable biomaterials in recent times; however, the requirement for optimized fibrillation conditions limits their practical use. Therefore, under standing the molecular mechanism of SSP amyloidogenesis, biochemical conditions, and the biophysical prop erties of the resultant amyloid fibrils becomes crucial. This study investigates the amyloidogenic properties of Cruciferin-3 (CRU-3), a major SSP from Arabidopsis thaliana, focusing on the 12-residue representative pep tide, L 223–Y 234 (LY12), computationally predicted to be amyloidogenic. LY12 forms β-sheet rich amyloid fibrils in a nucleation-dependent manner in vitro with the potential to seed self-aggregation. The peptide fibrillation was found to be pH-dependent and showed a moderate resistance to Proteinase K treatment. Molecular-level insight into the structure of LY12 fibrils was obtained using cryogenic-electron microscopy (cryo-EM) at a high reso lution of 2.86 Å. The structure of LY12 fibrils revealed a C2 symmetrical, left-handed, twisted core comprising three non-equivalent peptide stacks. This unique cross β-sheet dense core, stabilized by hydrophobic and elec trostatic interactions, and surrounded by low-density peptide layers, distinguishes them from pathological am yloids. This study explores the conditions for LY12 amyloid formation and deciphers their biophysical attributes and structural details, suggesting the potential physiological roles and biomaterial applications of CRU-3 amyloids