During the second period, we have further developed the LTS and LRS materials. For this purpose, synthetic biology tools were developed and used, focussing on 3 main organisms (S. cerevisiae, K. rheaticus, and B. subtillis). Key proteins vital for Living Therapeutic Skin (LTS) structure (hydrophobins, ELPs, CBDs, SpyTag, and SpyCatcher domains) have been effectively secreted or surface-displayed by yeast. Significant progress has been made in the synthesis of hyaluronic acid and acetylated cellulose by K. rhaeticus with strain development now underway. For the Living Regenerative Skin (LRS) material, a B. subtilis strain was genetically engineered to express cellulose binding domains to the surface of bacterial spores.
We also worked on advancing biomolecular engineering in order to further develop bio-encapsulation strategies. This research led to the successful expression of three distinct bacterial enzymes within S. cerevisiae. To achieve this, we utilized an expression system built upon the developed Yeast Toolkit (YTK). To optimize protein expression, we implemented a high-throughput screening approach that incorporated a yeast display procedure. This method was executed within an inter-laboratory setup, leveraging the capabilities of fluorescence-activated cell sorting (FACS) technology. Furthermore, we successfully expressed silk proteins and conducted in-depth analyses of thin film formation at the air-water interface. Additionally, we explored and evaluated multiple techniques for enhancing the stability of these films when transferred onto a solid support.
Regarding materials fabrication, we successfully developed prototype versions of both the LTS and LRS materials. While these prototypes are not yet fully functional, they already exhibit several of the intended functional properties that were originally proposed, such as biosensing. These early-stage materials provide a valuable opportunity to demonstrate and assess some of the expected material characteristics, as well as certain key biological properties. This progress serves as an important step toward further refinement and optimization in future iterations. A range of different approaches for shaping materials was explored, including 3D printing. These investigations aimed to assess the feasibility and effectiveness of various fabrication methods in achieving the desired structural and functional properties of the materials. 3D printing and other shaping strategies are very useful as we evaluate the potential of the LTS and LRS in terms of scalability and suitability for specific applications. This work lays the foundation for optimizing material processing techniques to enhance their overall performance and utility.
To gain insight into how potential users might perceive and interact with the LTS material, we developed mimics of their anticipated tactile and responsive properties. These initial prototypes were incorporated into user studies with individuals diagnosed with atopic dematisis (AD) to collect feedback on their usability and sensory experience. Building on the insights gathered from these studies, we refined the prototypes further in preparation for a workshop with our Impact Panel members. During this session, we engaged in discussions about the practicality, appeal, and broader social implications of the proposed applications, aiming to evaluate their potential impact and future development.
In order to explore and evaluate the potential applications of the LRS material, we began by conducting an extensive review of both existing literature and practical implementations related to the use of biomineralization in design. This comprehensive research process allowed us to gain valuable insights into current advancements, challenges, and opportunities in the field. As a key outcome of this review, we developed a structured taxonomy that categorizes various approaches and methodologies associated with biomineralization. This taxonomy will serve as a foundational framework for systematically selecting and refining application ideas in the next phase of our project. Following this initial research phase, we organized a workshop aimed at identifying innovative and viable applications for the LRS material. This collaborative session brought together members from another project within the EIC ELM Portfolio, facilitating interdisciplinary discussions and knowledge exchange. By engaging with experts from different domains, we sought to uncover novel use cases, assess the feasibility of various concepts, and generate ideas of applications.
