Safety considerations
Despite the fact that S. elongatus UTEX 2973 is thought to be safe for both humans and the environment, it is still our responsibility to ensure that our modified strains will be kept within our laboratories to eliminate any possible harm that can occur due to leakage. To prevent the exchange of genes or plasmids between our modified strains and wild strains, the photobioreactors used will be kept isolated from the outside environment to ensure safety. The growing public awareness regarding biosafety and the environment also serves as a reminder to us to maintain the control of our work. Since S. elongatus UTEX 2973 or is not a common organism seen or heard in our daily lives, we will also teach the public and our end-users that our strain is harmless.
End-users
The major end-users of our project are anticipated to be those in the nylon industry and pharmaceutical industry. We envision them implementing our approach to the synthesis of valerolactam with S. elongatus UTEX 2973 as to replace the current petrochemical industrial methods, to lower the harm caused to the surrounding environment. However, as many people in Hong Kong or worldwide have never heard of synthetic biology, it is understandable if manufacturers are hesitant to employ our method. To address their worries, we performed assays such as western-blot gel electrophoresis and capturing of valerolactam with magnetic beads. This shows that they can depend on the quality and quantity of the valerolactam manufactured in our project.
Cultivation of S. elongatus UTEX 2973 with wastewater
As we hope to further contribute to protecting the local and global environment, we wish to seek ways to culture S. elongatus UTEX 2973 that minimize the use of additional chemicals or nutrients and promote the sustainability of our project. We envision a cultivation method that uses wastewater as the nutrient base for the culturing and growth of S. elongatus UTEX 2973, as wastewaters often contain rich amounts of nitrogen and phosphorus essential for the growth of cyanobacteria. As reported by Ziagova and Amanatidou, S. elongatus PCC 7942, another S. elongatus strain, can be cultivated in wastewater that has been disinfected by combining physical disinfection of filtration and chemical disinfection with Fe(VI) [1]. If the same cultivation process can be done with S. elongatus UTEX 2973, not only can it help the environment by reusing wastewater meant to be disposed, it can also allow manufacturers to lower their costs of valerolactam synthesis even with large-scale implementation of our project.
Magnetic beads to capture valerolactam
After the synthesis of L-lysine, 5AVA and then cyclizing it into valerolactam, we propose to use magnetic beads to perform protein-ligand fishing, with the assistance of the NphR3 protein. The NphR3 protein, which can attach to valerolactam will be binded to magnetic beads and added into the mixture after cyclization. A magnetic will then be used to attract the beads such that the valerolactam gets removed from the mixture. After that, the captured valerolactam will be dissolved and can undergo further purification, whereas the magnetic beads can be reused for capturing again. This proposed method allows manufacturers or end-users to scale up their production to meet industrial needs as the magnetic beads are reusable and can save costs needed for extracting valerolactam from the original mixture.
Reference:
- Ziagova, Maria G., and Elisavet Amanatidou. “Wastewater Substrate Disinfection for Application of Synechococcus Elongatus PCC 7942 Cultivation as Tertiary Treatment.” Research Sqaure, Dec. 2021, https://doi.org/10.21203/rs.3.rs-1067481/v1.