14-ERASynBio: Synthetic Glycobiology - new strategies to build and functionalise proto-cells and proto-tissues

Great advances have been made in the development of proto-cells based on giant unilamellar vesicles (GUVs). However, one essential functional element of all living cells still to be incorporated into such systems is a glycocalyx. This coating of complex carbohydrates extends up to 100 nm from the cell membrane and provides an adhesive layer that mediates interactions between different cell types, viruses and signalling molecules. In most cases, these interactions involve specific carbohydrate-binding proteins (lectins) which may be either soluble or membrane-bound. For example, fertilisation is initiated by a specific carbohydrate on the surface of the egg adhering to a specific lectin on the head of the sperm. Protein-carbohydrate interactions also mediate the endocytosis of many bacteria, viruses and bacterial toxins which stick to specific glycolipids on the cell membrane. Protein-carbohydrate interactions thus present a general strategy for enabling cell adhesion and cell entry. In this project we will design and create a modular toolbox of synthetic glcocalyx components and engineered lectins that will be attached to lipid membranes to enable reversible proto-cell adhesion and incorporated into virus-like particles to mediate proto-cell entry. The methodology will be exemplified through the construction of proto-cells that contain "proto-organelles" and the assembly and remodelling of "proto-tissues" in which multiple types of proto-cells are brought together in a pre-defined fashion to create more complex systems.

Great advances have been made in the development of proto-cells based on giant unilamellar vesicles (GUVs). However, one essential functional element of all living cells still to be incorporated into such systems is a glycocalyx. This coating of complex carbohydrates extends up to 100 nm from the cell membrane and provides an adhesive layer that mediates interactions between different cell types, viruses and signalling molecules. In most cases, these interactions involve specific carbohydrate-binding proteins (lectins) which may be either soluble or membrane-bound. For example, fertilisation is initiated by a specific carbohydrate on the surface of the egg adhering to a specific lectin on the head of the sperm. Protein-carbohydrate interactions also mediate the endocytosis of many bacteria, viruses and bacterial toxins which stick to specific glycolipids on the cell membrane. Protein-carbohydrate interactions thus present a general strategy for enabling cell adhesion and cell entry. In this project we will design and create a modular toolbox of synthetic glcocalyx components and engineered lectins that will be attached to lipid membranes to enable reversible proto-cell adhesion and incorporated into virus-like particles to mediate proto-cell entry. The methodology will be exemplified through the construction of proto-cells that contain "proto-organelles" and the assembly and remodelling of "proto-tissues" in which multiple types of proto-cells are brought together in a pre-defined fashion to create more complex systems.

In the longer term, the neolectins, superlectins, virus-like particles and lectin-modified proto-cells will provide tools for delivering macromolecular drugs in a cell-specific fashion. At present most of the biopharmaceuticals on the market recognise cell surface targets, but the neolectins etc, will provide a mechanism for delivery into cells by endocytosis, and potentially intracellular trafficking to defined organelles. This advance will expand the range of medical targets accessible to biopharmaceuticals, while helping to reduce the high doses necessary for novel treatments that already act intracellularly, e.g., exon-skipping RNA based drugs. Applicant Turnbull already collaborates with GSK on macromolecular drug delivery using lectins. He also leads the targeted delivery theme of the University of Leeds Pharmaceutical and Biopharmaceutical Industry Hub which fosters translational collaborations between academic and industrial partners. Furthermore, the involvement of applicant Wiltschi from the Austrian Centre of Industrial Biotechnology (ACIB) will provide another key pathway to achieving impact from our research. The ACIB has 30 company partners and will function as a hub for the dissemination of results to potential industrial partners and the translational development of products involving neolectins and superlectins developed in SYNGLYCTIS.
Through this proposal we will contribute to the training of skilled individuals (including 8 early stage researchers) who will gain expertise in design, engineering and characterisation of protein and lipid assemblies for synthetic biology applications. The provision of skilled workers who are empowered to contribute to the development of both academic and industrial synthetic biology will have long term societal benefit for Europe. Their work in this project and through the rest of their careers will lead to new diagnostic methods and clinical treatments that will improve the health and quality of life for millions of Europeans.