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Many of the beautiful and often complex nanoscale structures in nature come about through the

spontaneous assembly of a set of molecular building blocks. Nature’s success in using this strategy is an inspiration to scientists, but also a source of envy. Despite significant advances in recent years, it remains extremely challenging to design synthetic molecules that self-assemble to create nanostructures with predictable size and shape. Lack of a detailed understanding of the non-covalent forces (such as hydrogen bonding) that hold the self-assembled structures together, and lack of the ability to predict the shapes of the molecular building blocks reliably, gives research in this field more of a trial-and-error character than is often admitted. Thus, there is a case for using an alternative approach in which the demands on the design are minimized and some of the control is handed back to the molecules. On page 19 of this issue, Rein Ulijn and colleagues at the University of Manchester show how the forces that control the self-assembly process can also be harnessed to drive the synthesis of the very molecules that self assemble1.

The work by the Ulijn group makes use of the principles of dynamic combinatorial chemistry — a powerful technique that emerged in the mid-1990s for exploring the ways in which molecules can ‘recognize’ each other2. Dynamic combinatorial libraries (DCLs) are mixtures that form through the reversible combination of building blocks. The product distribution of such mixtures is governed by the thermodynamic stabilities of all the individual constituents. Therefore, any non-covalent interactions that occur will influence the composition of the mixture, favouring those molecules that are most efficient at interacting with other molecules in the mixture. So far this has mainly been exploited for the discovery of small-molecule ligands that bind to biomolecules and for the development of new synthetic receptors.

However, DCLs are inherently self-screening: any non-covalent interactions within the mixtures also affect their product distributions. If a particular constituent in a DCL happens to bind to itself, it will promote its own formation at the expense of the other

compounds in the system. Although all DCLs can in principle show such behaviour, the present study by Ulijn and co-workers is one

of the first clear demonstrations that DCLs are also a powerful tool for the discovery of new motifs of self-assembly and new self-assembled

Molecular self-asseMbly

Helping themselvesBy using reversible enzyme reactions involving short peptides, molecular synthesis can be controlled by the self-assembly of the resulting products.

sijbren otto

Thermolysin

Figure 1 | Enzyme-assisted self-assembly. Starting from a dipeptide (top left) and an Fmoc-protected amino acid (top right), the enzyme thermolysin mediates the formation of reversible peptide bonds, giving rise to an equilibrium mixture of peptides of different lengths (middle). Of the various products, the tripeptide assembles efficiently into fibres (bottom), thereby shifting the equilibrium between the different peptides towards its own formation.

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14 nature nanotechnology | VOL 4 | JANUARY 2009 | www.nature.com/naturenanotechnology

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materials. Thermodynamic control permits error correction during both the synthesis of building blocks and the assembly process, making the resulting materials less prone to defects in comparison with materials that are produced under kinetic control.

The system described by Ulijn — now at the University of Strathclyde — and his team is based on reversible amide bond formation, which is a reaction involving the formation of a covalent bond from a carbonyl carbon (C=O) to a nitrogen atom. The reaction is mediated by thermolysin, an enzyme that can catalyse both amide bond formation and breakdown. The demonstration of reversible amide chemistry is a valuable addition to the limited set of reversible covalent chemistries available, and although researchers have pursued reversible amide chemistry for some time3, until now no good experimental procedure has been available. The report by Ulijn and co-workers is an important step forward, allowing access to dynamic mixtures of peptides (amides derived from amino-acid molecules) and bringing dynamic libraries of proteins a step closer.

The team showed that starting from dipeptides and amino acids protected by a fluorenyl (Fmoc) group, the action of thermolysin gives rise to a dynamic mixture of peptides of different lengths, containing typically one to five amino-acid residues (see Fig. 1). When using phenylalanine or lysine as the starting amino acid, the corresponding trimeric peptides were observed to self-assemble into fibres through a combination of stacking between the hydrophobic fluorenyl groups and β-sheet type main-chain hydrogen bonding. Not only do these interactions stabilize the nanostructures, they also cause the equilibrium between the various possible peptide products to shift in favour of the formation of the trimers. The fibres were of sufficient length and strength to give rise to entanglement and subsequent gelation of the aqueous solvent. Spatial control over the gelation process was also possible: when thermolysin was covalently immobilized on parts of a surface, gelation took place only in these locations.

This work demonstrates how interconnected chemical systems can

autonomously self-organize and give rise to new emergent behaviour. Here the action of an enzyme on a small set of very simple starting materials at the molecular level results in changes at the macroscopic level (gelation of the solvent). The coupling between gelation and the synthesis of the molecules that self-assemble into the fibres constituting the gel is essential for the new macroscopic behaviour to emerge. The success of this work will undoubtedly spark further activity in engineering interconnected chemical systems in search of new functions, and the opportunities arising from molecular networks are boundless, as nature demonstrates every day. ❐

Sijbren Otto is at the Centre for Systems Chemistry of the Stratingh Institute for Chemistry of the University of Groningen, 9747 AG Groningen, The Netherlands. e-mail: s.otto@rug.nl

references1. Williams, R. J. et al. Nature Nanotech. 4, 19–24 (2008).2. Corbett, P. T. et al. Chem. Rev. 106, 3652–3711 (2006).3. Bell, C. M., Kissounko, D. A., Gellman, S. H. & Stahl, S. S.

Angew. Chem. Int. Edn 46, 761–763 (2007).

Nanometre-scale particles derived from natural protein assemblies are increasingly being used to generate

new materials for diverse applications. These protein particles can exist as rods or spheres and can be used as templates for constructing more complex nanomaterials. They can also be chemically modified with a variety of functional groups or be laden with cargos in their interiors. Writing in Advanced Functional Materials, Nicole Steinmetz of the Scripps Research Institute and co-workers from the Institut Pasteur and the John Innes Centre1 report the chemical functionalization of a thermostable virus obtained from a microbe isolated from an acidic hot spring in Iceland. The ability to label these particles selectively with diverse chemical groups offers additional building blocks that can be used in a variety of harsh conditions, such as acidic environments in the body or the high temperatures found in some electronic devices2.

The use of viral nanoparticles as building blocks is advantageous because they are small (nanometre-sized), can self-assemble into discrete shapes and sizes, are highly symmetrical and polyvalent (that is, they have many binding sites) and can be produced easily in large quantities. Because the interior dimensions of the particles vary between 20 and several hundred nanometres and the genetic material can be removed, these protein cages can be rendered non-infectious and can be renovated to transport therapeutic payloads or to establish nanoscale reaction vessels. Among viruses that have shared the spotlight are the cowpea mosaic virus, which has been developed for applications in electronics and medicine, and the rod-shaped tobacco mosaic virus, which has been used in templates for making batteries and nanowires.

Altering the chemical properties of the virus particles has relied mainly on modifying surface-exposed amino acids naturally present in the viral coat proteins (particularly

the reactive groups present in lysine, cysteine, aspartate and glutamate). Attempts to install multiple modifications on virus particles have been hampered by the limited repertoire of conjugation reactions. Recently, significant advances have been made in both selective modification and robust assembly (Fig. 1). One approach is to exploit the intrinsic reactivity of other natural amino acids such as tyrosine (Fig. 1a). Matthew Francis and co-workers at the University of California, Berkeley, for example, selectively coupled these tyrosine residues to diazonium salts, which offers higher selectivity and modification efficiencies3. Tyrosine modification within the context of sequential conjugation strategies has also permitted the selective modification of the interior and exterior of both spherical and rod-like viral nanoparticles4,5 (Fig. 1b). It is also possible to enhance the number of different reactive groups on viral surfaces by introducing non-natural amino acids into coat proteins,

NaNoparticles

Designer labels for virus coatsProtein nanoparticles derived from viruses are commonly studied, but a new rod-shaped thermophilic virus isolated from acidic hot springs may yield another class of protein building blocks that are stable and can be selectively modified with diverse chemical groups.

isaac s. carrico and Kent Kirshenbaum

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