The project

The main objective of BIORE2AIRCRAFT is to develop a novel enzymatic-based method to depolymerise organic coating and use the degradation products in the manufacturing of new polymers, offering a cleaner and energy-efficient alternative to the current aerospace industry. This bigger aim can be divided into the following specific objectives:

(1) To bioinformatically and computationally screen and design novel enzymes with potential to degrade the organic polymers and composite materials found in aircraft fuel tanks based on a built knowledge graph of all enzyme sequence
(2) To design and optimise enzyme cascade reactions to depolymerise epoxy polymers
(3) To genomically edit Escherichia coli into an epoxy polymeric materials-degrading bacterium by incorporating the most promising enzyme cascade reactions, using CRISPR techniques
(4) To develop a technology that allows recovering the raw materials at high quality for aerospace applications and revalorisation of degradation products as softeners or liquid polymers for the manufacturing of new polymer materials enabling the circularity in aerospace

BIORE2AIRCRAFT involves an interdisciplinary methodology to discover novel enzymes to degrade the main epoxy polymeric and polyurethane materials’ components in the metallic parts of aircrafts. Metagenomic samples from aircraft fuel tanks microbiomes will be thoroughly analysed using the more recent computational tools. This will guide to the exploration of novel biochemical pathways capable of facilitating novel organic polymers depolymerisation reactions, as well as materialise protein sequences that assume completely new folds that diverge significantly from those documented in the record of natural evolution, and execute specific biochemical tasks. Throughout the exploration of the extremophilic microbiomes thriving in aircraft fuel tanks and related extreme microbiomes and generation of sequences, we will select and filter the functionally-driven generated sequences. A list of the most promising organic polymers-degrading enzymes will be obtained, and the likelihood to be expressed in E. coli will be assessed. The experimental characterisation of the selected enzymes will consist on trying them in the defined cascade reactions to see their performance against organic coating polymers. Checking their expression and catalytic activity towards the degradation of selected organic coatings components will be required. These enzymes will have to be gear parts of the defined enzyme cascade reaction for organic coating depolymerisation. Once the degradation pathway for organic coating is established a novel synthetic biology approach will be proposed.

This will be achieved in three ways. Firstly, by co-expressing the needed enzymes in a plasmid, in such a way that successful extracellular expression is ensured. Second, by designing genomically CRISPR-edited E. coli strains that integrates the enzyme candidates involved in the cascade reaction into its genome, enabling the bacteria to produce them extracellularly. Thirly, leveraging comprehensive knowledge of the structures and active sites of these enzymes, designing a genomically CRISPR-edited E. coli equipped with its own extracellular proteins evolved for the degradation of organic coating compounds in polymers is proposed. E. coli has been chosen as the model chassis because synthetic biology tools are available. Once the evolved and pre-adapted strain of E. coli with the incorporated genes is obtained, the next steps are to conduct its fermentation in the presence of the epoxy polymer material, and carry out downstream processing steps to separate the degradation products. Depending on the degradation level of the organic part different approaches are possible: direct use into the chemical industry is possible and the best possible chance for circular economy, or used as a plasticisers directly for the manufacturing of new polymer materials. Degradation products will be prepared in various concentrations and qualities, then sent to our collaborator, Airbus, for evaluating product properties. Standard trials for manufacturing new polymer materials for aerospace applications, enabling circularity, will be conducted.
In this way, BIORE2AIRCRAFT’s technological advancements and approach will result in ready-to-use genomically edited E. coli strains for the treatment of raw metallic structures of aircrafts containing polyurethane resins blended with epoxy resins without any previous pretreatment, and their reuse.