Bioinspired control of protein transport through polymer functionalised nanopores

This project will initiate a research programme in nanopore protein separation that is inspired by the nuclear pore complex (NPC).

NPCs are complex, giant assemblies constituted from more than 400 proteins that define nanoscale pores (i.e. nanopores) ~40 nm in diameter. Each NPC spans the nuclear membrane which separates the cell nucleus and the rest of the cell. NPCs are the only conduits in and out of the cell nucleus in all eukaryotic cells and they allow only a small set of specific proteins and genetic material related to the functioning of the cell nucleus to pass through. All the other thousands of species of unrelated but similar molecules in the cell are rejected.

Convenient separation of biomolecules is an enabling technology. NPC-studded nuclear membranes are effectively a highly specific and efficient molecular separation and purification membrane. They are capable of sorting through more than 1 kg of specific biomolecules in a human body per minute, far surpassing the performance of current technology. The creation of NPC-mimetic nanoporous membranes would benefit diverse biotechnology and biomedical applications, ranging from purification of protein disease markers for bedside medical diagnosis to continuous manufacturing of enzymes and protein therapeutics. Understanding the science underlying NPC function will help us achieve these applications and help us meet our 21st century challenges in healthcare and advanced manufacturing.

The immediate goal of this project is to establish the design rules for enabling the basic function of the NPC - the sorting of proteins according to size using nanopores with a "virtual" size cut off and which, unlike current technology, are not clogged by random interactions with proteins. The pore size of the NPC is virtual because it has a physical diameter much larger than the size of the protein. A random protein cannot however pass through because each NPC nanopore is filled with a semi-porous polymer plug with an as yet unidentified structure that specifically repels proteins, except for those proteins specific to nuclear function.

Biologists studying the NPC have proposed two leading theories to explain how the plug works: i) the "virtual gate" polymer brush model, and ii) the
"selective phase" meshwork model. This project will create artificial nanopores that are decorated with synthetic polymers as simplified models to mimic these two theoretical structures. Experiments will be conducted to verify whether either of the theories is in fact feasible.

The ultimate goal is to exploit these design rules for further development of the nanoporous membrane platform that incorporate increasingly advanced polymers for decorating the nanopores. This will create NPC-inspired nanoporous membranes with separation efficiency and selectivity that matches, and may eventually even surpass, native NPC function.

Who are the users of the research and how will they benefit?
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ACADEMIC and INDUSTRY RESEARCH USERS, as well as the specific benefits accrued to them, are identified in the section "Academic beneficiaries". Multiple disciplines will benefit, including polymer science, nanomaterials, biointerfaces, and structural biology.

COMMERCIAL ENTERPRISES engaged in the above research areas, as well as those interested in protein separation and nanoporous membrane technologies will also be direct users. They will benefit from the new knowledge generated to create a separation technology that is significantly more functional than the state-of-the-art in terms of molecular specificity, operating life, throughput, and flow pressure and energy requirements. Importantly, the membrane format is highly scalable - the membrane area can be selected to suit either lab-on-a-chip or manufacturing scales.

THE WIDER PUBLIC will be the end-users and eventual beneficiaries. Convenient separation of biomolecules, especially proteins, will be transformative. The benefits are aligned with the EPSRC's Prosperity Outcomes in Productivity and Health. Overall, the technology is enabling in its capabilities and disruptive in its scalability, and will engender new business and technological opportunities. New and improved applications in biomedicine and biotechnology will improve the quality of life. Improvements to the continuous manufacturing of protein therapeutics and enzymes will lower the costs to medical treatment and high-value chemical products. Rapid and low cost biosensing as well as new capabilities in drug delivery may be enabled to improve and widen access to healthcare. (See Pathways to Impact for details.)

Furthermore, this research is aligned with goals identified by the EPSRC to enhance the UK economy. The broad field of users illustrates the INTERDISCIPLINARY NATURE of the research. The project will initiate a CUTTING-EDGE INTERNATIONAL COLLABORATION with Prof Théato, Head of the Institute for Technical and Macromolecular Chemistry (ITMC) at the University of Hamburg. Also a PDRA will be trained in state-of-the-art research skills to DELIVER THE HIGHLY NUMERATE MANPOWER the UK needs.

What will be done to ensure that users will benefit?
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The project results will be disseminated through scientific journals and conferences targeted for their high international profile and scientific relevance.

The PI will undertake industrial visits to keep up to date with industry needs and to generate interest in the technology being developed. The visits will include Meissner Filtration Products Inc., USA, which has expressed an interest (see Letter of Support), and corporations invested in the areas of nanoporous membranes and molecular separation (e.g. Millipore, GE Healthcare).

In addition, a workshop on "Polymer Surfaces And Functional Nanoporous Membranes" will be organised for the academic and industry users to exchange research results and application ideas, and forge R&D opportunities.

To explain the benefits to the public, a public presentation at the Glasgow Science Centre (GSC) will be held, and the PI's website will advertise the results and impact, and Strathclyde's public relations office will issue press releases.

The PDRA will be trained through regular PI supervision as well as through the budgeted dissemination activities (presentation at conference and GSC). Moreover, a 2-week training visit to the Théato group at Hamburg is budgeted.