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Project Description

Protein Nanocompartments

Protein nanocompartments (PNCs) have the ability to encapsulate, deliver and help integrate cargos into various systems. Using PNCs with targeting abilities with the use of surface peptides not only makes delivery of cargos more specific but also increase efficiency. This can also limit off targeting effects that may come with cargos such as therapeutics [REF}. Therefore due to its simple use and broad applicability PNCs are a valuable tool not only for future iGEM teams but for the scientific community as well.

Applications

Drug Delivery: due to the potential for specific targeting with the addition of surface peptides, PNCs are excellent contenders for enhancing drug delivery and therefore effectiveness of said drug. For example, increasing targeting and delivery of chemotherapeutics to affect only cancer cells with a lower dosage [REF].
Targeted Transformations/Transfections of Cell Populations: the ability for PNCs to deliver and integrate cargos into cell membranes gives this system the potential to provide easier and more efficient ways of incorporating plasmid DNA or other materials into cell populations. Including bacterial cells and more perverse systems.
Targeted Antibiotic Delivery
Vaccine Development
Gene Therapy
Materials Synthesis

Our System

Our goal for the 2018 iGEM season is to create a PNC toolkit for future iGEM teams. A software tool will be created to help individuals build PNCs that cater to their specific project needs which include surface peptides, encapsulation proteins and specific cargo loading approaches that can be used for various applications. With these designs we can address a wide range of issues such as antibiotic resistance, the negative side-effects of chemotherapeutics, or the impact of using GMOs as biological control agents. In the wetlab we will be addressing various applications with several virus-like capsids.

P22: The major capsid of the P22 bacteriophage will be used to encapsulate FitD, a molluscicidal toxin derived from Pseudomonas fluorescens.2 PNCs will be aggregated using a modified P22 decoration protein linker to encourage consumption by Zebra mussels in an effort to reduce invasive populations in Canadian waters.
MS2: To demonstrate targeted delivery of small molecules, the capsid protein from the MS2 bacteriophage is modified with a polyarginine surface peptide to encourage capsid incorporation into a cell.4 For proof-of-concept, we will demonstrate efficient encapsulation and delivery of green fluorescent protein (GFP) cargo. The GFP will be modified with a negatively-charged peptide to enhance packaging and eliminate the need for RNA-initiated capsid assembly.
Arc Gag: Similar to homologous viruses, this mammalian protein self-assembles to form capsid-like structures that shuttle Arc mRNA between cells.6 We will determine the recognition sequence for mRNA encapsidation and the minimal Arc protein required to form capsids for use in a simplified cell culture transfection procedure.

Biocontainment

By using PNCs instead of bacteria to deliver cargoes such as the FitD toxin, we prevent releasing modified bacterial populations into the environment and therefore bring a viable and safe alternative for eradicating non-indigenous mussel populations from affected areas. The purified protein components can prevent the release of recombinant DNA material and prevent any effects from potential uptake by other organisms found in the environment.

Biosecurity

We recognize the opportunity for misuse of the proposed software that enables users to design their own targeting PNCs and cargos technology. It is feasible that a user could include pathogenic nucleic acids or a human toxin as PNC cargo. To mitigate these implications, we will integrate an automated fail-safe into our software whereby the system determines if the cargo designed for the PNC is homologous to known dangerous sequences. In the current climate of biosafety regulations, we hope to encourage a sense of responsibility amongst potential users and promote the adoption of simple biosecurity measures wherever possible.

Human Practices

While virus-like particles are well documented in the literature and have demonstrated potential as vaccines, other real world applications of PNCs are currently limited. Thus, we will also engage current experts in the healthcare field to explore the feasibility of PNC use in gene therapy, drug delivery, and more.

References

Protein nanocompartments (PNCs) have the ability to encapsulate, deliver and help integrate cargos into various systems. Using PNCs with targeting abilities through the use of surface peptides not only makes delivery of cargos more specific but also increases efficiency. This can also limit off targeting effects that may come with cargos such as therapeutics. Therefore, due to its simple use and broad applicability, PNCs are a valuable tool not only for future iGEM teams but for the scientific community as well. Our goal for the 2018 iGEM season is to create a PNC toolkit for future iGEM teams. With these designs we can address a wide range of issues such as antibiotic resistance, the negative side-effects of chemotherapeutics, or the impact of using GMOs as biological control agents. One problem that we are focusing on is the presence of the invasive zebra and quagga mussels in bodies of water. We plan to build on the use of the sequencing technologies we used for our 2016 project to detect the presence of the mussels in water and then to use the P22 capsid with the toxin FitD to prevent further spread of the mussels. We will also be using the viral capsids MS2 and Arc Gag to demonstrate the efficacy of our design for small molecule transport and cell culture transfection, respectively. We plan on engaging stakeholders in the environmental and health care fields in addition to scientific researchers to ensure that our project is able to meet the needs of multiple user groups.

We are also developing a software tool to help individuals build PNCs that cater to their specific project needs. The software will allow the user to input components such as surface peptides, encapsulation proteins and specific cargo loading approaches that can be used for various applications. We recognize the opportunity for misuse of the proposed software that enables users to design their own targeting PNCs.To mitigate these implications, we will integrate an automated fail-safe into our software whereby the system determines if the cargo designed for the PNC is homologous to known dangerous sequences. In the current climate of biosafety regulations, we hope to encourage a sense of responsibility amongst potential users and promote the adoption of simple biosecurity measures wherever possible.