Contribution

Beyond addressing a pressing medical need, our project also serves the broader iGEM community by contributing valuable biological tools and insights. These resources can aid future iGEM teams as they pursue projects that, like ours, aim to benefit society through synthetic biology innovations.

A Lack of Suitable Parts and Information

When we began this project, it was apparent that there was a significant lack of literature, information, and available composite parts in the iGEM Parts Registry related to synthetic biology research using B. subtilis. Specifically, finding an effective method to secrete recombinant enzymes from B. subtilis was particularly challenging, as few resources existed to guide us. We aimed to fill these gaps and provide a foundation for future teams working in human enzyme-related fields, specificially in B. subtilis.

Help from other iGEM Teams

In developing our system, we utilized 11 existing parts from the iGEM Parts Registry. By applying these parts in new contexts, our work will broaden the community’s understanding of how these parts function, particularly when integrated into complex biological systems in other chassis, such as B. subtilis. Our data will enhance the collective knowledge surrounding these parts, and help future teams incorporate them into their own designs more effectively.

Our Contribution to the Registry

Alongside leveraging existing tools, we contributed five new parts to the iGEM registry, expanding the resources available for future synthetic biology projects. The first part is the human pancreatic lipase protein encoding gene (PNLIP) with a signal peptide optimized for protein secretion in Bacillus subtilis. As this signal peptide is a novel feature of our project, we hope this serves as a model or idea for future iGEM teams working toward systems permitting secretion of enzymes. Another one of our parts is the human procolipase protein encoding gene (CLPS), also containing a signal peptide optimized for protein secretion in B. subtilis. Our parts related to the glucose-mediated switch mechanism include amyE cre sites that enable inducible glucose expression via the carbon catabolite repressible pathway, and an RNA anti-terminator (RAT) element that provides gene repression within promoter regions. These parts are also novel and we hope future iGEM teams take advantage of this part when designing inducible systems for gene expression.

Composite Parts

Our team assembled these parts into unique composite parts, each with a specific function. These include a composite part for lipase production and secretion and a composite part for procolipase production and secretion. We also developed a composite part for inducible glucose expression, which can be regulated based on external glucose conditions. Finally, we created a biocontainment-focused composite part designed to disable the entire device if unsatisfactory conditions are detected, preventing potential environmental repercussions if the engineered bacteria leave the human body or enter unintended environments. These composite parts demonstrate a cohesive, well-rounded approach to enzyme therapy design and safety.

Future Directions and Remaining Needs

While we have made significant progress, our experimentation is still incomplete. In the near future, we aim to further develop and test our device to determine the yield of enzymes secreted by the device. We plan to incorporate post-translational modifications, stronger promoters, more extensive biocontainment, and other key components to maximize efficiency and applicability of our device.

Furthermore, we intend to explore different variations of signal peptides and map their kinetics to determine the optimal sequences for the secretion of recombinant enzymes from B. subtilis. Understanding how these peptides affect enzyme secretion will not only improve our project but will also offer valuable insights to the wider iGEM community working on recombinant enzyme production for therapeutic applications.

By contributing to the iGEM database, we aim to establish a solid foundation for future teams to build upon. We encourage others to explore, expand, and refine our work, potentially driving innovations in enzyme therapy or probiotics, particularly for pancreatic and other enzyme-deficiency disorders. The availability of our sequences, protocols, and findings will allow future teams to improve enzyme stability, increase yields, or develop more efficient delivery methods.

Beyond our research goals pertaining to device development, we aimed to expand the reach of synthetic biology. To promote inclusivity and accessibility to this rapidly-growing field, we developed a synthetic biology handbook for the homeschooled community. Our 40-page handbook contains engaging content such as DIY-experiments, activities, and other interactive materials for students to learn about synthetic biology. The handbook provides a brief and exciting introduction to the field, with the intention of igniting a deeper passion for this field in young students. Though our handbook is primarily geared towards homeschooled students between the ages of 11 and 17 years old, it will be made accessible and available for any individuals interested in learning more about synthetic biology. Our handbook aims to eliminate all barriers to this field and promote education of synthetic biology to a broader community than ever before.