Toward the development of peptide nanofilaments and nanoropes as smart materials

Abstract

Protein design studies using coiled coils have illustrated the potential of engineering simple peptides to self-associate into polymers and networks. Although basic aspects of self-assembly in protein systems have been demonstrated, it remains a major challenge to create materials whose large-scale structures are well determined from design of local protein-protein interactions. Here, we show the design and characterization of a helical peptide, which uses phased hydrophobic interactions to drive assembly into nanofilaments and fibrils (''nanoropes''). Using the hydrophobic effect to drive self-assembly circumvents problems of uncontrolled self-assembly seen in previous approaches that used electrostatics as a mode for self-assembly. The nanostructures designed here are characterized by biophysical methods including analytical ultracentrifugation, dynamic light scattering, and circular dichroism to measure their solution properties, and atomic force microscopy to study their behavior on surfaces. Additionally, the assembly of such structures can be predictably regulated by using various environmental factors, such as pH, salt, other molecular crowding reagents, and specifically designed ''capping'' peptides. This ability to regulate self-assembly is a critical feature in creating smart peptide biomaterials.biomaterial ͉ coiled coil ͉ protein design ͉ circular dichroism ͉ atomic force microscopy D esigned proteins are valuable paradigms for engineering at the nanoscale, offering favorable properties such as atomiclevel precision and tight regulation of self-assembly by using a variety of environmental cues (i.e., pH, ionic strength, temperature) (1-3). As one of the most well studied and naturally abundant structural motifs in proteins, coiled coils are particularly suited to protein design. Their basic sequence feature, the heptad repeat, is a seven-residue pattern (abcdefg) n of nonpolar and polar residues that gives rise to amphipathic ␣-helices. The hydrophobic effect drives the burial of nonpolar residues at the helix-pairing interface and influences geometric details of resulting structures (4). Electrostatic and polar residues at buried or solvent-exposed locations provide additional means to manipulate structural features by stabilization or destabilization (negative design) of key interactions (5-8).Recent attempts to design self-assembling protein networks by using coiled coils have focused on simple systems, such as linear and branched fibrils (9-14), planar assemblies (15), and hydrogels (16-18). Specifically, filament formation has been achieved by stabilization of intermediate structures that foster intermolecular coiled-coil interactions (11)(12)(13)(14). Previous studies have reported the design of short helical peptides, which interact via a dimeric coiled-coil motif and are stabilized by a combination of overlapping hydrophobic and electrostatic intermolecular interactions (9, 13, 14). These peptides do indeed self-assemble into filaments; however, these filaments also show evidence of extensive...

Engineering nanoscale order into a designed protein fiber

“…Predominantly, such work has used ␤-structured peptides that form amyloid-like structures (3, 5, 6, 10). Relatively less has been attempted with ␣-helix-based assemblies, however (7,(11)(12)(13)(14). The development of ␣-helical systems would provide useful comparisons with the more-explored amyloid-like systems and also allows the application of the considerable body of protein design and engineering knowledge for ␣-helical structures and assemblies (15)(16)(17).…”

Self-assembly of synthetic collagen triple helices

“…Using the self-assembly approach, Woolfson and coworkers (18,19) have produced fibers with a design based on a dimeric coiled-coil structure. Likewise, fibrous peptides based on the natural protein elastin (20) and de novo building blocks (15,(21)(22)(23)(24)(25)(26) have been assembled and implicated as biomaterial candidates. Here, we report on the self-assembly of collagen fragments into an extended trimeric coiled-coil.…”

Crystal structure of a super leucine zipper, an extended two‐stranded super long coiled coil

“…In another nanorope assembly system, 21 the self-assembling leucine zipper peptide has an insertion of two alanine residues at the middle of the helix, and as a result of the insertion the N-and C-terminal hydrophobic ridges are on the two opposite faces of the helix. In the proposed nanorope assembly model, 21 the C-terminal hydrophobic ridge of the first helix is attached to the N-terminal hydrophobic ridge of the second helix, whereas the C-terminal hydrophobic ridge of the second helix is attached to the N-terminal hydrophobic ridge of the third helix. The overall nanorope assembly elongates in a staggered mode, in which the helices are offset by half length of the helix or $2 heptad.…”