Triangular in Vivo Self-Assembling Coiled-Coil Protein Origami

Abstract

Coiled-coil protein origami (CCPO) polyhedra are designed self-assembling nanostructures constructed from coiled coil (CC)-forming modules connected into a single chain. For testing new CCPO building modules, simpler polyhedra could be used that should maintain most features relevant to larger scaffolds. We show the design and characterization of nanoscale single-chain triangles, composed of six concatenated parallel CC dimer-forming segments connected by flexible linker peptides. The polypeptides self-assembled in bacteria in agreement with the design, and the shape of the polypeptides was confirmed with small-angle X-ray scattering. Fusion with split-fluorescent protein domains was used as a functional assay in bacteria, based on the discrimination between the correctly folded and misfolded nanoscale triangles comprising correct, mismatched, or truncated modules. This strategy was used to evaluate the optimal size of linkers between CC segments which comprised eight amino acid residues.

Coiled-coil protein origami nanostructure modeling for improved characterization and prediction

“…We further performed modeling of two distinct triangular CCPO structures (TRI6 (ref. 28) and DoNA-GN 4 (ref. 34)) that contain different linkers of (SG) 4 or (GN) 4 , respectively (Fig.…”

“…Additionally, structure models of two distinct triangular CCPO structures (TRI6 (ref. 28) and DoNA-GN 4 (ref. 34)) were built, following the same procedure through 10 cycles of MD optimization.…”

“…37 Using known coiled-coil structures as templates, comparative models of various CCPO designs have been built and fitted to SAXS data. 26–29,32…”

Peptide Design and Self-assembly into Targeted Nanostructure and Functional Materials

“…10 nm in size using a single oligopeptide chain that folded into six pairs of homodimeric coiled coils (Figure ). They later showed protein-like folding of peptides in bacteria into closed triangular loops held together by peptide dimers and flexible linkers . Woolfson and co-workers have constructed stochastic self-assembling cage particles or “SAGE” particles that are ca.…”

Metal ion–regulated assembly of designed modular protein cages

“…By correctly combining CC-forming peptides or linking them to other oligomerizing units, diverse CC-based nanostructures can be created (9)(10)(11). We have previously shown that CC dimers can be used to design mono-and multimeric protein cages of different polyhedral shapes (15)(16)(17)(18)(19). This strategy, termed CC protein origami (CCPO), is based on the concatenation of orthogonal CC-forming peptides (20) into one or several polypeptide chains that self-assemble into the desired fold determined by the pattern of pairwise interactions between the CC-forming modules.…”