BIOMOLECULE-DIRECTED EVOLUTION OF INORGANIC NANOMATERIALS

From the extremely simple micro-organisms which appeared billions of years ago, the diversity and complexity of the plants and animals currently surrounding us results from evolution. In turn, the ability of organisms to evolve is itself attributed to the DNA which forms the blueprint of life on earth. DNA can undergo mutations, some of which may generate a "stronger" organism. As organisms exist in a competitive environment, the stronger species survive to transmit their "winning" DNA to new progeny and future generations.

In this interdisciplinary research project, we gain inspiration from nature to use DNA-based technologies to evolve inorganic nanomaterials with targeted properties. With the production of structures such as bones, teeth and seashells, nature shows that it is possible to produce inorganic materials whose properties are perfectly optimised for their function under very mild conditions. While organisms clearly achieve this using many strategies, all are united by one common feature; nature controls the formation of inorganic solids using organic molecules. These biomolecules are themselves the result of evolutionary selection such that the fittest survive to produce materials with target size, shape, orientation and polymorph.

Expertise in molecular biology has now reached levels where DNA technologies can be used to evolve huge libraries of biomolecules. In combination with high throughput screening methods, it has therefore become possible to generate biomolecules for increasingly diverse target applications. While this exciting reduction of biological evolution to the laboratory timescale has been used to improve applications such as organic catalysis, its huge potential in materials synthesis remains almost entirely untapped. This research proposal will address this challenge, and employ a novel approach to evolve DNA-encoded nanomaterials. Our strategy is based on three key factors.

(1) We will utilize two diverse DNA libraries - which encode for libraries of biomolecules - that have never before been screened for material synthesis, but that are very well suited for this purpose.

(2) We will utilize a completely new micro-droplet-based platform (using microfluidic devices). Single DNA molecules encapsulated within single micro-droplets will be used to express unique biomolecules. Nanoparticles of cadmium sulfide, copper sulfide and magnetite (magnetic iron oxide) will then be synthesised within these unique micro-environments.

(3) We will screen directly for the PROPERTIES of the synthesized nanomaterials. Droplets containing "winning" nanoparticles with target photoluminescent or magnetic properties will be isolated using fluorescence activated cell sorting (FACS) or magnetic separation. Recovery of the DNA from the "winning" droplets then enables expression of the "winning" protein, which can be employed for large-scale synthesis of the winning nanoparticles.

Finally, our experimental approach will enable us to link biomolecule structure and function. While researchers have for the last 50 years studied biomolecules extracted from biominerals, we still have a very poor understanding of how these control factors such as polymorphism. Here, our diverse libraries are derived from a unique protein scaffold ("adhiron") that exhibits the desirable property of being readily crystallized. Profiting from this ability, we will determine the 3D structure of "winning" proteins, and in comparison with selected "losers", will be able to gain unique insight into the origin of their activity.

This proposal will use directed-evolution techniques to synthesise inorganic materials with target properties. Through this strategy we will lay the groundwork for materials scientists to readily combine DNA technologies and high throughput screening methods to identify and generate biomolecules which are able to support the formation of materials with specific properties. Importantly, our methods will also deliver environmentally-friendly and sustainable processing routes which operate in aqueous solutions at ambient temperature. Currently, methods to identify organic molecules capable of controlling the formation of inorganic materials are largely emprical, being based on trial-and-error.

The beneficiaries of this research programme therefore include academic and industrial researchers working in the field of inorganic materials synthesis, which includes areas such as nanotechnology, energy and catalysis. The methods developed can also be used to control, for example, crystallization phenomena. These are fundamental to technological processes and natural phenomena as diverse as the production of pharmaceuticals, foodstuffs and personal care products, the formation of biominerals, the precipitation of ice in the atmosphere and the prevention of scale. It will also impact on those working in synthetic biology, which has been highlighted as an area of strategic importance by the EPSRC. There, it is expected that biological systems, or key components of biological systems will have the potential to generate new products and processes for UK industries. By linking synthetic biology to materials technologies we will identify key proteins capable of generating nanomaterials with specific properties.

The topics listed above serve to demonstrate that the project has the potential to make a significant impact on the wider public, and also on the UK economy, in areas ranging from healthcare to everyday technologies. Indeed, within the time-scale of the project we would expect to have identified biomolecules capable, for example, of controlling the synthesis of magnetite in aqueous solution. Magnetite is widely used in biomedical applications, but, while highly uniform particles have been synthesised at high temperatures in organic solvents, it has not yet been possible to achieve this in aqueous solution. The widespread adoption of the methods developed here for identification of biomolecules capable of generating inorganic materials with specific properties is likely to go beyond the lifetime of this project. However, we would expect our approaches to inspire other researchers to link synthetic biology with materials synthesis, making our goals achievable in the near future.

This project will also provide the postdoctoral researchers with opportunities to acquire a wide range of skills. Both PDRAs will work closely throughout the project, enabling them to gain complementary research skills. Opportunities in areas such as writing papers, giving presentations at conferences, networking with other researchers, exploring possible patent opportunities, attending staff development courses and taking part in outreach work will provide excellent chances for professional development.

Finally, the wider public will also benefit through a programme of outreach work, in which all of the researchers will participate. Leeds hosts the annual "Leeds Festival of Science" where local schools take part in educational workshops related to science and engineering, and we will design a workshop relating to the evolution of materials, and its relevance to everyday life. We will also prepare a "Schools Talk" based on this material. In addition, the investigators will continue to run and facilitate a Leeds iGEM synthetic biology team of undergraduate students, which will enhance their scientific and team working skills, and they will deliver widening participation activities during the projects.