Japan_IPAP: Embedding protein crystals within synthetic tissues as catalytic soft materials
Lead Research Organisation:
Imperial College London
Department Name: Chemistry
Abstract
Bottom-up synthetic biology aims to reproduce the structures and behaviours of cellular organisms by combining molecules that mimic the structural, functional and information containing roles present in biology. These structures are known as synthetic cells, and they can act as a framework to study biological processes (such as movement or replication), as well as act as miniature test-tubes, within which chemical and biochemical reactions can be carried out. Taking further inspiration from biology, the assembly of individual synthetic cells leads to the creation of synthetic tissues, where different compartment types can be integrated within a larger structure to act as optimised microscale reaction vessels, which can be activated through the exchange of molecular reactants between compartments.
As synthetic cells and tissues are constructed from molecular parts, there is huge flexibility in the design of these systems, and this has been exploited to form compartments from lipids, polymers and protein-based membranes. One area that is significantly less explored is the encapsulation of novel biocatalysts structures within synthetic cells and tissues. One such catalyst are polyhedrin protein crystals, which can be used to embed co-produced enzyme catalysts via expression of crystals in cells. This exploits the high stability of polyhedrin crystals (used to protect viral capsids in the external environment in nature) to create long-lasting catalysts.
Here, we propose to use microfluidic approaches to assembling synthetic tissues that i) encapsulate enzyme-embedded polyhedrin crystals within different compartments of the tissue and ii) possess a hydrogel shell to increase the durability of the tissue material. We will assemble water in oil emulsion droplets on-chip, encapsulating the crystals within the aqueous compartments of these droplets, before gelating the hydrogel around the emulsion to produce milliscale devices that can be handled in liquid and air. We will then test the catalytic activity of these tissues, with the aim of producing robust soft materials with long catalytic lifetime that can be used simply via incubation in reactant-containing solution. After reactant takeup and conversion within the synthetic tissue, we will aim to release product through washing cycles, setting up the next tissue-based catalysis cycle. This proof-of-concept work will demonstrate the potential for hybrid tissue mimics in catalysis, and this framework could be extended to design new, combined bio- and chemo-catalytic routes currently impossible in one-pot systems due to catalyst poisoning.
In order to achieve this we are bringing together world leading research expertise in the UK and Japan with a view to bringing together expertise not available in concert elsewhere in the word: namely Ces (synthetic cells and microfluidics), Ueno (multiscale catalytic biomaterials) and Abe (catalytic protein crystal biomaterials).
As synthetic cells and tissues are constructed from molecular parts, there is huge flexibility in the design of these systems, and this has been exploited to form compartments from lipids, polymers and protein-based membranes. One area that is significantly less explored is the encapsulation of novel biocatalysts structures within synthetic cells and tissues. One such catalyst are polyhedrin protein crystals, which can be used to embed co-produced enzyme catalysts via expression of crystals in cells. This exploits the high stability of polyhedrin crystals (used to protect viral capsids in the external environment in nature) to create long-lasting catalysts.
Here, we propose to use microfluidic approaches to assembling synthetic tissues that i) encapsulate enzyme-embedded polyhedrin crystals within different compartments of the tissue and ii) possess a hydrogel shell to increase the durability of the tissue material. We will assemble water in oil emulsion droplets on-chip, encapsulating the crystals within the aqueous compartments of these droplets, before gelating the hydrogel around the emulsion to produce milliscale devices that can be handled in liquid and air. We will then test the catalytic activity of these tissues, with the aim of producing robust soft materials with long catalytic lifetime that can be used simply via incubation in reactant-containing solution. After reactant takeup and conversion within the synthetic tissue, we will aim to release product through washing cycles, setting up the next tissue-based catalysis cycle. This proof-of-concept work will demonstrate the potential for hybrid tissue mimics in catalysis, and this framework could be extended to design new, combined bio- and chemo-catalytic routes currently impossible in one-pot systems due to catalyst poisoning.
In order to achieve this we are bringing together world leading research expertise in the UK and Japan with a view to bringing together expertise not available in concert elsewhere in the word: namely Ces (synthetic cells and microfluidics), Ueno (multiscale catalytic biomaterials) and Abe (catalytic protein crystal biomaterials).
Technical Summary
Bottom-up synthetic biology aims to construct lifelike systems and materials from their constitutive molecular parts (e.g. lipids, proteins). This has led to the engineering of synthetic cells (unicellular) as well as tissue-like structures (multi-cellular) that exhibit the ability to communicate with their surroundings and undertake multi-step synthesis, exploiting compartmentalisation to generate multiple chemical environments in the same structure.
As synthetic cells and tissues are constructed from molecular parts, a plug-and-play approach is facilitated for cell design, enabling the assembly of pathways and the use of molecular assemblies not found in nature. One unexplored area is the use of novel catalysts. The compartmentalisation properties of synthetic cells and tissues facilitate the integration with novel biocatalytic materials, for example ultrastable polyhedrin protein crystals that can embed bio (e.g. enzymatic) or chem (e.g. metal) catalysts. Encapsulation within the crystal structure extends enzyme durability and lifetime, facilitating their use in industrial biocatalysis.
Here, we propose to integrate enzyme-containing protein crystals into synthetic cells based on droplet-based, hydrogel-coated emulsions which are assembled into synthetic tissues using microfluidics. Single crystal-based enzyme catalysts are isolated in a tissue compartment, enabling the assembly of orthogonal microreactors with optimised reaction conditions within the same material. We will use this concept to build a two-step crystal-localised reaction pathway through the compartments of a synthetic tissue, acting as proof-of-concept for the design of more complex tissue-based catalysts in future UK and Japanese funded projects. This could unlock catalytic soft materials containing new bio and chemo-catalysed synthetic routes currently impossible to assemble in currently used one-pot processes, leading to the application of synthetic tissues in industrial biocatalysis.
As synthetic cells and tissues are constructed from molecular parts, a plug-and-play approach is facilitated for cell design, enabling the assembly of pathways and the use of molecular assemblies not found in nature. One unexplored area is the use of novel catalysts. The compartmentalisation properties of synthetic cells and tissues facilitate the integration with novel biocatalytic materials, for example ultrastable polyhedrin protein crystals that can embed bio (e.g. enzymatic) or chem (e.g. metal) catalysts. Encapsulation within the crystal structure extends enzyme durability and lifetime, facilitating their use in industrial biocatalysis.
Here, we propose to integrate enzyme-containing protein crystals into synthetic cells based on droplet-based, hydrogel-coated emulsions which are assembled into synthetic tissues using microfluidics. Single crystal-based enzyme catalysts are isolated in a tissue compartment, enabling the assembly of orthogonal microreactors with optimised reaction conditions within the same material. We will use this concept to build a two-step crystal-localised reaction pathway through the compartments of a synthetic tissue, acting as proof-of-concept for the design of more complex tissue-based catalysts in future UK and Japanese funded projects. This could unlock catalytic soft materials containing new bio and chemo-catalysed synthetic routes currently impossible to assemble in currently used one-pot processes, leading to the application of synthetic tissues in industrial biocatalysis.
People |
ORCID iD |
Oscar Ces (Principal Investigator) |
Description | Collaboration with Takafumi Ueno (Tokyo Institute of Technology) |
Organisation | Tokyo Institute of Technology |
Country | Japan |
Sector | Academic/University |
PI Contribution | To employ our expertise in soft matter, microfluidics and our synthetic cell models as chemical compartments to host protein crystal catalysts. |
Collaborator Contribution | To employ their expertise in the development of protein crystal catalysts which serve as the functional molecule in this project. The Ueno laboratory produced and characterised the crystal catalysts. |
Impact | We are currently applying for follow-up funding through the co-funded BBSRC/JST Japan-UK engineering biology for discovery research and cross-cutting technologies with Takafumi Ueno and other researchers at Tokyo Institute of Technology. |
Start Year | 2023 |