Engineering synthetic pathways to bio-ethylene production in Cyanobacteria sp.
Lead Research Organisation:
University of Nottingham
Department Name: School of Life Sciences
Abstract
Background: Ethylene is currently produced from the stream cracking of ethane which produces large quantities of CO2, contributing to global warming. In 2000, steam cracking had a primary energy use of 3 billion Gigajoules and accounted for approximately 200 million tons of CO2 emissions. Ethylene is the monomer for the most common plastic, polyethylene, and annual global production is approximately 80 million tons. Finding a sustainable or carbon neutral alternative to ethylene production is imperative. Cyanobacteria are Gram-negative photosynthetic prokaryotes. Researchers have previously attempted to produce ethylene from CO2 through heterologous expression of the efe gene in the cyanobacteria, Synechococcus elongates sp. PCC 7942 and Synechocystis sp. PCC 6803, with mixed results.
Aim: The aim of this project is to engineer cyanobacteria as a platform for the production of hydrocarbon-based products such as ethylene. Expression of the efe gene encoding the ethylene-forming enzyme from Pseudomonas syringae pv. Phaseolicola in Synechocystis sp. PCC 6803, led to continuous ethylene production. We are now in the process of improving production through directed evolution and metabolic engineering. As part of this process we would like to engineer a synthetic pathway for ethylene production utilising the Yang pathway from plants. This provides an exciting opportunity to implement a novel pathway in cyanobacteria and link ethylene production to growth. Ethylene is efficiently biosynthesized from 1-aminocyclopropane-1-carboxylic acid (ACC) (Zhou et al., 2002), which is itself derived from methionine as a branch of the Yang cycle (Wang et al., 2002). This process is energetically efficient as it preserves the high-energy methionine thioether bond. This pathway utilises SAM synthetase, ACC synthase and ACC oxidase. The conversion of 1-amino-cyclopropane-1-carboxylic acid (ACC) to ethylene releases cyanoformic acid, which spontaneously decarboxylates to release hydrogen cyanide, which is principally detoxified by the CAS pathway (Machingura et al., 2016). The implementation of this pathway will provide a mechanism for detoxifying cyanide in cyanobacteria.
Training: The project will allow for training in a unique multidisciplinary environment, incorporating genomic engineering, gas fermentation, synthetic biology, cutting edge molecular biology and systems biology modelling. The project will providing the student with a vast array of transferable skills, highly prized by employers in the growing bio-economy. The project will also provide several high impact publications. This translational project will be carried out within the BBSRC/EPSRC Synthetic Biology Research Centre (SBRC) at Nottingham which comprises 70+ graduate and postdoctoral researchers (www.clostron.com/people.php) and a current budget of £27M.
Aim: The aim of this project is to engineer cyanobacteria as a platform for the production of hydrocarbon-based products such as ethylene. Expression of the efe gene encoding the ethylene-forming enzyme from Pseudomonas syringae pv. Phaseolicola in Synechocystis sp. PCC 6803, led to continuous ethylene production. We are now in the process of improving production through directed evolution and metabolic engineering. As part of this process we would like to engineer a synthetic pathway for ethylene production utilising the Yang pathway from plants. This provides an exciting opportunity to implement a novel pathway in cyanobacteria and link ethylene production to growth. Ethylene is efficiently biosynthesized from 1-aminocyclopropane-1-carboxylic acid (ACC) (Zhou et al., 2002), which is itself derived from methionine as a branch of the Yang cycle (Wang et al., 2002). This process is energetically efficient as it preserves the high-energy methionine thioether bond. This pathway utilises SAM synthetase, ACC synthase and ACC oxidase. The conversion of 1-amino-cyclopropane-1-carboxylic acid (ACC) to ethylene releases cyanoformic acid, which spontaneously decarboxylates to release hydrogen cyanide, which is principally detoxified by the CAS pathway (Machingura et al., 2016). The implementation of this pathway will provide a mechanism for detoxifying cyanide in cyanobacteria.
Training: The project will allow for training in a unique multidisciplinary environment, incorporating genomic engineering, gas fermentation, synthetic biology, cutting edge molecular biology and systems biology modelling. The project will providing the student with a vast array of transferable skills, highly prized by employers in the growing bio-economy. The project will also provide several high impact publications. This translational project will be carried out within the BBSRC/EPSRC Synthetic Biology Research Centre (SBRC) at Nottingham which comprises 70+ graduate and postdoctoral researchers (www.clostron.com/people.php) and a current budget of £27M.
Organisations
Studentship Projects
| Project Reference | Relationship | Related To | Start | End | Student Name |
|---|---|---|---|---|---|
| BB/M008770/1 | 01/10/2015 | 31/10/2024 | |||
| 1797253 | Studentship | BB/M008770/1 | 01/10/2016 | 17/02/2021 |
| Description | In order to generate ethylene by heterologous expression of the plant pathway/Yang cycle gene in E. coli both genes - ACC Synthase (ACS) and ACC oxidase (ACO) must be correctly transcribed and translated into active protein. So far wild type/non codon optimised genes have been used. Plant enzymes can often pose problems when expressed in prokaryotes, due to rare codons, cleavage sites and post-transnational modifications. Thus far aco1 from Solanum lycopersicum has been expressed and identified by rtPCR and western blot. acs1 from Malus domestica however, has only been detectable by rtPCR, and no active protein has been detectable. The result has been, no detectable ethylene production in strains harbouring both acs1 and aco1. Codon optimised genes have now been employed and results will hopefully follow shortly. |
| Exploitation Route | Any work with Malus domestica acs1 in prokaryotic hosts must be carried out with either fully codon optimisation, or at least codon optimisation of the first 9 N-terminal amino acids as in White et. al 1994 (PMID: 7809054). |
| Sectors | Manufacturing, including Industrial Biotechology |