There are several approaches to creating synthetic-biological systems. Here, we describe a molecular-design approach. First, we lay out a possible synthetic-biology space, which we define with a plot of complexity of components versus divergence from nature. In this scheme, there are basic units, which range from natural amino acids to totally synthetic small molecules. These are linked together to form programmable tectons, for example, amphipathic alpha-helices. In turn, tectons can interact to give self-assembled units, which can combine and organize further to produce functional assemblies and systems. To illustrate one path through this vast landscape, we focus on protein engineering and design. We describe how, for certain protein-folding motifs, polypeptide chains can be instructed to fold. These folds can be combined to give structured complexes, and function can be incorporated through computational design. Finally, we describe how protein-based systems may be encapsulated to control and investigate their functions.

Small peptides offer an attractive starting point for the development of self-assembling materials for a variety of purposes, since they are relatively simple to produce and can be tailored to provide an expansive range of chemical functionality. We have employed a short peptide that spontaneously self-assembles into a multimolecular fibrillar architecture to drive the coassembly of two independent luminescent moieties. We use fluorescence spectroscopy to demonstrate that the resulting complex performs a light-harvesting function.

Osmotic shock in a vesicle or cell is the stress build-up and subsequent rupture of the phospholipid membrane that occurs when a relatively high concentration of salt is unable to cross the membrane and instead an inflow of water alleviates the salt concentration gradient. This is a well-known failure mechanism for cells and vesicles (for example, hypotonic shock) and metal alloys (for example, hydrogen embrittlement). We propose the concept of collective osmotic shock, whereby a coordinated explosive fracture resulting from multiplexing the singular effects of osmotic shock at discrete sites within an ordered material results in regular bicontinuous structures. The concept is demonstrated here using self-assembled block copolymer micelles, yet it is applicable to organized heterogeneous materials where a minority component can be selectively degraded and solvated whilst ensconced in a matrix capable of plastic deformation. We discuss the application of these self-supported, perforated multilayer materials in photonics, nanofiltration and optoelectronics.

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We describe the formation of self-assembling nanoscale fibrillar aggregates from a hybrid system comprising a short polypeptide conjugated to the fluorophore fluorene. The fibrils are typically unbranched, approximately 7 nm in diameter, and many microns in length. A range of techniques are used to demonstrate that the spectroscopic nature of the fluorophore is significantly altered in the fibrillar environment. Time-resolved fluorescence spectroscopy reveals changes in the guest fluorophore, consistent with energy migration and excimer formation within the fibrils. We thus demonstrate the use of self-assembling peptides to drive the assembly of a guest moiety, in which novel characteristics are observed as a consequence. We suggest that this method could be used to drive the assembly of a wide range of guests, offering the development of a variety of useful, smart nanomaterials that are able to self-assemble in a controllable and robust fashion.

We describe a straightforward single-peptide design that self-assembles into extended and thickened nano-to-mesoscale fibers of remarkable stability and order. The basic chassis of the design is the well-understood dimeric alpha-helical coiled-coil motif. As such, the peptide has a heptad sequence repeat, abcdefg , with isoleucine and leucine residues at the a and d sites to ensure dimerization. In addition, to direct staggered assembly of peptides and to foster fibrillogenesisthat is, as opposed to blunt-ended discrete speciesthe terminal quarters of the peptide are cationic and the central half anionic with lysine and glutamate, respectively, at core-flanking e and g positions. This +,-,-,+ arrangement gives the peptide its name, MagicWand (MW). As judged by circular dichroism (CD) spectra, MW assembles to alpha-helical structures in the sub-micromolar range and above. The thermal unfolding of MW is reversible with a melting temperature >70 degrees C at 100 muM peptide concentration. Negative-stain transmission electron microscopy (TEM) of MW assemblies reveals stiff, straight, fibrous rods that extended for tens of microns. Moreover, different stains highlight considerable order both perpendicular and parallel to the fiber long axis. The dimensions of these features are consistent with bundles of long, straight coiled alpha-helical coiled coils with their axes aligned parallel to the long axis of the fibers. The fiber thickening indicates inter-coiled-coil interactions. Mutagenesis of the outer surface of the peptide i.e., at the b and f positionscombined with stability and microscopy measurements, highlights the role of electrostatic and cation-pi interactions in driving fiber formation, stability and thickening. These findings are discussed in the context of the growing number of self-assembling peptide-based fibrous systems.

Interest in the design of peptide-based fibrous materials is growing because it opens possibilities to explore fundamental aspects of peptide self-assembly and to exploit the resulting structures--for example, as scaffolds for tissue engineering. Here we investigate the assembly pathway of self-assembling fibers, a rationally designed alpha-helical coiled-coil system comprising two peptides that assemble on mixing. The dimensions spanned by the peptides and final structures (nanometers to micrometers), and the timescale over which folding and assembly occur (seconds to hours), necessitate a multi-technique approach employing spectroscopy, analytical ultracentrifugation, electron and light microscopy, and protein design to produce a physical model. We show that fibers form via a nucleation and growth mechanism. The two peptides combine rapidly (in less than seconds) to form sticky ended, partly helical heterodimers. A lag phase follows, on the order of tens of minutes, and is concentration-dependent. The critical nucleus comprises six to eight partially folded dimers. Growth is then linear in dimers, and subsequent fiber growth occurs in hours through both elongation and thickening. At later times (several hours), fibers grow predominantly through elongation. This kinetic, biomolecular description of the folding-and-assembly process allows the self-assembling fiber system to be manipulated and controlled, which we demonstrate through seeding experiments to obtain different distributions of fiber lengths. This study and the resulting mechanism we propose provide a potential route to achieving temporal control of functional fibers with future applications in biotechnology and nanoscale science and technology.

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