Functional Biomolecular Liquids

Designing new materials that have small-scale (nanometre) structures and combine multiple components is expected to lead to the development of new technology in areas of sensing, electronics, catalysis and medicine. These materials can be difficult to synthesise, and a new approach is to use large biological molecules known as proteins as an active component. Proteins are made up of long chains of amino acids that fold upon themselves to form complex 3D structures. In the body, proteins perform a wide variety of tasks (or functions) from the binding of oxygen in muscles (which is performed by the protein myoglobin), to the storage of iron in the blood (ferritin), and it would be advantageous if these properties could be transferred to a synthetic material. Proteins are most commonly found either as dispersions in aqueous solutions or as dry powders, and it is fascinating to note that till recently, proteins in the pure liquid phase did not exist, i.e. heating a dry protein powder will not cause it to melt. In essence, this means that there was a missing phase of biological matter that was yet to be discovered.
The absence of a pure liquid protein phase results from the relatively large molecular dimensions (nanoscale) of the protein molecule, and is an intriguing phenomenon that is also seen with nanoparticles. The situation arises because the liquid phase of a material is stabilized by attractive inter-molecular forces that act over distances that are considerably larger than the size of the individual molecules. This is not the case for proteins however, as their structures are large compared with the range of the forces between them. In essence, the protein molecules are so firmly held together in the solid phase that heating would not make them melt, but rather, would destroy their molecular structure, resulting in decomposition.
The aim of my research is to design a universal approach to access the missing liquid phase of proteins by increasing the range of the attractive protein-protein interactions. To do this I will attach artificial (synthetic) polymer surfactant molecules to the proteins' surfaces to produce protein molecules with long tendrils that can interact with other protein molecules over longer distances. These polymer surfactant molecules are negatively charged, and only attach to positively charged groups on the protein surface. Hence it will be necessary to first chemically alter the surface of the protein molecules to make them more positively charged, so that enough of the polymer surfactant molecules can be attached. In my preliminary studies I used this approach to produce liquids of ferritin and myoglobin, which contained no water and melted near room temperature. What was truly astounding is that even though the protein molecules have evolved to operate in aqueous environments, their structures in the pure liquid phase appeared not to have changed, and in the case of myoglobin, the protein could still bind oxygen.
My proposed work allows me to apply my knowledge of biochemistry, materials science and physical chemistry to develop a new class of hybrid biological liquids, and I intend to develop this new approach to produce a wide range of liquid proteins with different functions. In each case I will investigate the molecular structure of the liquids, as well as their composition and properties such as viscosity, and I will also test the protein for function. This will not only provide a range of new active liquids, but will aid in the understanding of how important water is for protein structure and function. Finally, once I understand how these systems work, then I will use the results to develop new types of materials based on liquid proteins. For example, I intend to develop new biological sensors for the detection of toxic gases such as carbon monoxide, or active wound dressings that supply oxygen to the wound during healing.

Scientific innovation is essential for the international competitiveness of the UK, and bionanotechnology will play a key role in the advancement of new technologies that will have significant impact on national economies. Within this expansive field, the development of new classes of biological materials and the associated methods of synthesis of these bio-nanomaterials, will provide new directions for scientists working at the interfaces of chemistry, molecular biology, medicine and materials science. Through the development of a library of functional biomolecular liquids, this project will deliver an unprecedented advance in the EPSRC's Physical Sciences Capability and Challenge Theme priority area of Chemical Biology and Biological Chemistry, while providing vital information that will further our fundamental understanding of the role of solvent molecules in protein structure, dynamics and function.

The proposed research describes a method to extend the phase diagram of proteins to include a solvent-free liquid phase, which will increase the general utility and practical applications of proteins in science and technology. The development of these materials is expected to impact on a diverse range of disciplines including biochemistry, chemistry and medicine, and contribute to the UK's competitiveness and economic performance through the development of new biomaterials and devices. Functional biomolecular liquids contain protein concentrations far in excess of the aqueous solubility limit, and accordingly will provide new avenues for the application of industrial enzymes such as lipases and proteases in the detergent industry; the development of new wound dressings (glucose oxidase liquids) for the healthcare industry; and the effective storage of protein-based pharmaceuticals. The development of membrane-active protein-polymer surfactant conjugates will also impact on healthcare technologies via improvements in the efficiency of tissue engineering, and through the detoxification of organophosphates using phosphotriesterase biomolecular liquids. Recently, AWP had confidential discussions with Dr. Michael Butler of Unilever on application of functional biomolecular liquids for new enzyme-based detergent formulations. Moreover, there has been significant interest in our preliminary work on the use of these materials for tissue engineering, and we are presently in discussions with the University of Bristol's Research Enterprise and Development (RED) office about securing a provisional patent. We plan to continue to participate in the science innovation training provided by RED to fully maximise impact on the commercial sector over the duration of the research programme.

The new science generated by the discovery and exploration of these novel materials will be published in top-tier peer-reviewed journals, which will act as vehicles for the dissemination of knowledge to potential beneficiaries. AWP's recent work on ferritin and myoglobin liquid proteins has attracted much publicity and has been featured in a number of applied engineering and pharmaceutical journals, as well as in the popular science media (see Case for Support, Part 1). This will be complemented by participation at international conferences in order to communicate the potential technology to interested parties in the private sector. We anticipate that this research will also impact internationally upon policy-makers and international funding bodies by demonstrating the value of chemical biology research. This research will also be promoted within the University of Bristol via the Research News website, and this will raise the university's profile within the UK and allow us to establish new international links and collaborations across multiple departments and disciplines.