What is the impact of our project?
Enzymes and reagents for molecular biology, the means of biotechnological production, are expensive and hard to access, especially for people in developing countries and outside academia and industry. This problem has been acutely felt through enzyme shortages during the COVID-19 pandemic. The wetware supply chain fails miserably in many parts of the world.
Inexpensive access to the enzymes required for biotechnological research and development, combined with low-cost hardware like nanopore DNA sequencers, robotic liquid handlers, plate readers and bioreactors, could mean that high-capacity biofoundries (for the assembly and testing of genetic devices from DNA parts, as well as the manufacturing of biological products, such as enzymes) are now within reach, even for resource-constrained biotechnologists.
Integrated together, these pieces of hardware, software, wetware and lab-ware could constitute a new and largely open-source biotechnological infrastructure stack (biostack) that greatly lowers the financial and technical barriers to bioengineering. In turn, this could rapidly enable greater numbers of bioengineers, and hubs of biotechnological research and manufacturing, around the world. This is critically important, because the accelerating climate crisis demands that humanity both transform the material base of its civilization to sustainable modes of production, and re-shape our manufacturing processes and supply chains to make communities everywhere resilient to disruptive climate shocks.
Our goal is to design and assemble a Frugal Enzyme Production and Purification Process. In particular, we aim to build genetic devices that induce the bacteria Bacillus subtilis and the yeast Pichia pastoris to efficiently secrete our target enzymes (including Phusion DNAP, T4 ligase and polynucleotide kinase, and Type IIS restriction enzymes like BsaI and BtgzI), as this secretion will greatly simplify downstream processing and purification. Because it is difficult to rationally design constructs with efficient and high-yield protein secretion, we will also attempt to integrate various low-cost laboratory hardware, including an Opentrons liquid handler, a nanopore DNA sequencer, and an open source plate reader, into a frugal biofoundry that enables us to build and test many different constructs in parallel. Once we express and secrete proteins, we will establish a protocol pipeline for the purification of enzymes using the most scalable and inexpensive approaches possible (i.e. ELP tag separation, cosmotropic salt precipitation, and frugal chromatography with silica-binding peptide tags).