Team:Brasil-USP/Project/Overview
Overview
Project
Overview
Brasil-USP 2015 team intends to apply synthetic biology to solve a serious environmental and social problem: waste tires accumulation. Developing such project is interesting once current used recycling approaches are inefficient, demanding large amounts of energy and resulting in low yields of low quality subproducts. Furthermore, waste tires are frequently associated with tropical diseases dissemination and may take up to 1000 years until complete natural degradation. That being said, our project aims the construction of an entire process for tire recycling, generating a final product with commercial value: a biofuel. The whole procedure is divided into three different steps: devulcanization, biodegradation and chemical transformation (Figure 1). The first step is destined to treat the tire and revert rubber vulcanization, a process needed to confer hardness and resistance to natural rubber so it can be used in the tire manufacturing procedure. The devulcanization is naturally done by a microorganism, Acidithiobacillus ferrooxidans, that is capable of removing the sulfur added to the natural rubber during vulcanization1. The main product of this first step is natural rubber, which is the main substrate for the second step, that consists in its proper treatment with our genetically modified organism. The design of efficient rubber degradation bacteria (link pro chassis: http://parts.igem.org/Escherichia_coli_chassis) includes the co-expression of two enzymes: RoxA (Rubber oxygenase A) and Lcp (Latex clearing protein). Both RoxA and Lcp were recently shown to successfully catalyze rubber cleavage to multiple smaller products2,3. To extracellular export RoxA and Lcp, we added 5’ signals to their coding sequence, while the biosafety is accomplished by an effective kill switch system based on HokD expression under a TetR inducible promoter. The output of this stage is a mix of different molecules, but the most important and abundant one is called 12-oxo-4,8-dimethyl-trideca-4,8-diene-1-al (ODTD), a tri-isoprene unit4. This molecule, with few chemical modifications, can be turned into a biofuel suitable for airplane use, our final product.
We intend to scale-up this project using bioreactors, but we still need further studies to make this possible. Modeling of every step is being made together with the experiments whenever possible Intending to evaluate the reality surrounding tire disposal in our country, we visited the National Association of Pneumatic Industry (ANIP), a non-profit organization that represents pneumatic industries in Brazil; CBL Reciclagem, a brazilian rubber recycling company and Recur Environmental Solutions, a company that aims to market by-products from tire pyrolysis. Theses visits, besides enriching our experience around the subject, got us in touch with the current technologies used in rubber recycling and waste tires management, making clear the need for new technologies to deal with this problem, fully availing our project. To know more about each aspect of our project, please click on the links below!
References
1 Kanao T.; Kamimura K.;, Sugio T. “Identification of a gene encoding a tetrathionate hydrolase in Acidithiobacillus ferrooxidans”; J Biotechnology, 132(1), 16-22, 2007 Oct 21. 2 Braaz, R.; Fischer, P. and Jendrossek, D. “Novel Type of Heme-Dependent Oxygenase Catalyzes Oxidative Cleavage of Rubber (Poly-cis-1,4-Isoprene)”. Appl. Environmental Microbiology, 70 (12), 7388-7395 (2004). 3 Hiessl, S., Böse, D., Oetermann, S., Eggers, J., Pietruszka, J., Steinbüchel, A. Latex Clearing Protein—an Oxygenase Cleaving Poly(cis-1,4-Isoprene) Rubber at the cis Double Bonds. Kivisaar M, ed. Applied and Environmental Microbiology. 80(17), 5231-5240 (2014). 4 Birke, J., Jendrossek, D. Rubber oxygenase and latex clearing protein cleave rubber to different products and use different cleavage mechanisms. Applied and Environmental Microbiology, 80 (16), 5012-5020 (2014).