Contributions

The bioreactor design

Solid state bioreactor model
The destruction of Halocarbons is a festering problem. While we tried to engineer a microbial, sustainable way to address this, we agreed that it will not be a solution if we do not have a device to employ the microbe. Our bioreactor design went through thorough multiple cycles of engineering, with a financial and efficiency analysis of every model. Our final solid state model with some novel caveats is pitch-perfect for degrading halocarbons and similar compounds.

3D render of solid state bioreactor

Parts

We used a fairly uncommon chassis, Pseudomonas putida, well-known for its applications in bioremediation and biotechnology. Along with our parts for the enzyme systems Cytochrome P450cam and Toluene dioxygenase, we worked on characterizing an ANR-regulated hypoxic promoter native to Pseudomonas, that can deactivate the downstream genes under aerobic conditions. In addition, we added documentation to the registry part BBa_K3765010 that may prove useful to future teams intending to use this part for a similar purpose.

Chemical Assays

We did not have access to facilities like a headspace analyzer that are generally used to quantify the rates of halocarbon degradation reactions. Through a series of discussions with Prof. Wackett and Maddy Bygd from the University of Minnesota, and the faculty at IISc, we designed and characterized a modified version of the general protocol. We improved it as we experimented and troubleshooted and used it to check the basal bacterial degradation rates of halocarbons.

Molecular Docking

We used molecular docking methods to test the binding of halocarbon substrates and intermediates on our enzymes compared to their cognate substrates. The results helped us identify the most probable binding sites and the possible reaction pathway. Through the in-silico tests for a set of gases, we could finalize the ones that should be used first for chemical assays and development of a proof of concept. We have characterized the docking protocol for two immensely diverse enzymes native to Pseudomonas, Cytochrome P450cam and Toluene dioxygenase. A comprehensible enzyme kinetic model of these enzymes were simulated adding information to the existing literature.

Molecular Docking

Docking of R-142b at Cytochrome P450cam active site

Educational Package

Over the months, we have developed a robust educational package that caters to students, and people across all ages. Special focus was put on reaching out to the rural students and our synthetic biology booklet, 'Sync', was translated in 10 languages that constitute the first tongue for 50% of the globe. Its content was improvised through critical analyses by both educators and people from the community, and was highly appreciated by our collaborating iGEM teams. To improve accessibility, we got the material translated into sign language. Find more details here!