Microbial conversion of lignin to monomers for bio-based plastics using synthetic biology (MILIMO)
Lead Research Organisation:
University of Warwick
Department Name: Chemistry
Abstract
There is a societal need in EU countries to develop new types of bio-plastics that could be used to replace current petroleum-based plastics for a range of packaging applications. The industrial partner in this Project, Biome Bioplastics Ltd (UK), has been developing new polyester plastics containing replacements for terephthalic acid used in petroleum-based plastics such as PET or PBAT. Professor Bugg and Biome Bioplastics have developed a biocatalytic route from the biopolymer lignin, found in plant cell walls, to 2,4- and 2,5-pyridinedicarboxylic acid, which can be used as biobased replacements for terephthalic acid. This project will seek to enhance the yields of 2,4-PDCA and 2,5-PDCA from lignin, to the point where a commercially feasible method of production could be developed.
The project will use synthetic biology methods to enhance the rate of lignin oxidation in two lignin-degrading bacteria: Rhodococcus jostii RHA1 and Pseudomonas putida KT2440. We will also aim to convert hydroxycinnamic acids or the cellulose fraction of lignocellulose into PDCA monomers. Synthetic biology will also be used to redesign the metabolic pathways of Pseudomonas putida KT2440 in order to optimise the production of PDCA monomers from different plant feedstocks.
We will also investigate the use of consortia of different lignocelluose-degrading bacteria for the conversion of lignin into PDCA monomers, mimicking the situation in soil where consortia of lignin-degrading specialists and cellulose-degrading specialists co-operate in order to break down lignocellulosic biomass.
Having developed a high-yield process for production of PDCAs on a small scale under laboratory conditions, the biocatalytic process will be scaled up to yield an efficient fermentation-based method for conversion of lignin or lignocellulose feedstocks to PDCA monomers, that could be commercialised by Biome Bioplastics.
The project will use synthetic biology methods to enhance the rate of lignin oxidation in two lignin-degrading bacteria: Rhodococcus jostii RHA1 and Pseudomonas putida KT2440. We will also aim to convert hydroxycinnamic acids or the cellulose fraction of lignocellulose into PDCA monomers. Synthetic biology will also be used to redesign the metabolic pathways of Pseudomonas putida KT2440 in order to optimise the production of PDCA monomers from different plant feedstocks.
We will also investigate the use of consortia of different lignocelluose-degrading bacteria for the conversion of lignin into PDCA monomers, mimicking the situation in soil where consortia of lignin-degrading specialists and cellulose-degrading specialists co-operate in order to break down lignocellulosic biomass.
Having developed a high-yield process for production of PDCAs on a small scale under laboratory conditions, the biocatalytic process will be scaled up to yield an efficient fermentation-based method for conversion of lignin or lignocellulose feedstocks to PDCA monomers, that could be commercialised by Biome Bioplastics.
Technical Summary
This project will seek to enhance the yields of 2,4- and 2,5-pyridinedicarboxylic acids (PDCA) from lignin, to the point where a commercially feasible method of production could be developed.
The project will use synthetic biology methods to enhance the rate of lignin oxidation in two lignin-degrading bacteria: Rhodococcus jostii RHA1 and Pseudomonas putidaKT2440. Enhancements in PDCA yield will also be sought via conversion of hydroxycinnamic acids or the cellulose fraction of pretreated lignocellulose into PDCA monomers. Synthetic biology will also be used to redesign the metabolic pathways of Pseudomonas putidaKT2440 in order to optimise flux from G- or S-type lignin along specialised pathways to PDCA monomers.
Microbial consortia will also be investigated for the conversion of lignin into PDCA monomers, mimicking the situation in soil where consortia of lignin-degrading specialists and cellulose-degrading specialists co-operate in order to break down lignocellulosic biomass. Hence the engineered G- and S-specialist degraders will be co-cultured with lignin-degrading L-specialists, or with other lignin-degrading micro-organisms. Specialist cellulose degraders will also be co-cultured in order to release glucose that could also be channelled to PDCA monomers by our engineered microbes.
Having developed high-yielding processes on a small scale under laboratory conditions, the biocatalytic process will be scaled up to yield an efficient fermentation-based method for conversion of lignin or lignocellulose feedstocks to PDCA monomers, that could be commercialised by Biome Bioplastics.
The project will use synthetic biology methods to enhance the rate of lignin oxidation in two lignin-degrading bacteria: Rhodococcus jostii RHA1 and Pseudomonas putidaKT2440. Enhancements in PDCA yield will also be sought via conversion of hydroxycinnamic acids or the cellulose fraction of pretreated lignocellulose into PDCA monomers. Synthetic biology will also be used to redesign the metabolic pathways of Pseudomonas putidaKT2440 in order to optimise flux from G- or S-type lignin along specialised pathways to PDCA monomers.
Microbial consortia will also be investigated for the conversion of lignin into PDCA monomers, mimicking the situation in soil where consortia of lignin-degrading specialists and cellulose-degrading specialists co-operate in order to break down lignocellulosic biomass. Hence the engineered G- and S-specialist degraders will be co-cultured with lignin-degrading L-specialists, or with other lignin-degrading micro-organisms. Specialist cellulose degraders will also be co-cultured in order to release glucose that could also be channelled to PDCA monomers by our engineered microbes.
Having developed high-yielding processes on a small scale under laboratory conditions, the biocatalytic process will be scaled up to yield an efficient fermentation-based method for conversion of lignin or lignocellulose feedstocks to PDCA monomers, that could be commercialised by Biome Bioplastics.
Planned Impact
The Project has an industrial partner (not eligible for funding under this Call), Biome Bioplastics Ltd, who are seeking to develop new bio-based plastics based upon PDCA monomers. The aim of this study is to develop a biocatalytic process for production of PDCAs from lignin or lignocellulose, that will then be commercialised by Biome Bioplastics Ltd. We therefore have a clear route to commercialisation for this work.
People |
ORCID iD |
| Timothy Bugg (Principal Investigator) |
Publications
Alruwaili A
(2023)
Elucidation of microbial lignin degradation pathways using synthetic isotope-labelled lignin.
in RSC chemical biology
Alruwaili A
(2023)
Application of Rhodococcus jostii RHA1 glycolate oxidase as an efficient accessory enzyme for lignin conversion by bacterial Dyp peroxidase enzymes.
in Green chemistry : an international journal and green chemistry resource : GC
Bugg T
(2021)
Microbial hosts for metabolic engineering of lignin bioconversion to renewable chemicals
in Renewable and Sustainable Energy Reviews
Spence EM
(2021)
Metabolic engineering of Rhodococcus jostii RHA1 for production of pyridine-dicarboxylic acids from lignin.
in Microbial cell factories
Yasin R
(2023)
Engineering of Rhodococcus jostii RHA1 for utilisation of carboxymethylcellulose
in Heliyon
| Description | Rhodococcus jostii RHA1 has been engineered to produce 2,4-pyridinedicarboxylic acid (2,4-PDCA) in improved yield by insertion of the ligAB genes onto the chromosome of this bacterium, and co-expression of a lignin-oxidising dypC gene. Overexpression of lignin oxidising genes from Rhodococcus jostii RHA1 and other lignin-degrading bacteria has identified two genes in Rhodococcus jostii RHA1 that, when overexpressed, lead to 2.7 and 3.5 fold enhancements in PDCA production. Rhodococcus jostii RHA1 has also been engineered to utilise cellulose as a carbon source, which will allow the conversion of both lignin and cellulose components of lignocellulose to PDCA bioplastic precursors. A new downstream processing method for high yield isolation of 2,4-PDCA from fermentation broth has also been developed, which will facilitate scale-up work on PDCA production. |
| Exploitation Route | Project partners Biome Bioplastics Ltd are interested in commercialising the results of this research, and have a current patent (from 2015) for the production of PDCA monomers from lignin feedstocks. |
| Sectors | Agriculture, Food and Drink,Chemicals,Manufacturing, including Industrial Biotechology |
| Description | Collaboration with Biome Bioplastics to generate monomers for bioplastic production from lignin degradation |
| Organisation | Biome Technologies plc |
| Country | United Kingdom |
| Sector | Private |
| PI Contribution | My research group has collaborated with Biome Bioplastics to develop a new process for generation of pyridine-dicarboxylic acids from lignin, by metabolic engineering of lignin degradation pathways in bacteria such as Rhodococcus putida and Pseudomonas putida. In 2015 we published the feasibility of this approach, which was also patented in 2015, and which has led to further collaborative projects. |
| Collaborator Contribution | Biome Bioplastics are interested in making new bio-based plastics from raw materials derived from lignin. They have brought prior knowledge in renewable bioplastics to the collaboration, and they hope to use the outputs of the project to develop a new process for production of a new bioplastic to the market. |
| Impact | Journal publications have been reported elsewhere in Research Fish. A patent application was also made in 2015. |
| Start Year | 2013 |
| Description | Collaboration with Dr Eduardo Diaz (CIB) to study of the molecular basis for lignin degradation in Pseudomonas putida |
| Organisation | Centre for Biological Research (CIB) |
| Country | Spain |
| Sector | Public |
| PI Contribution | We have collaborated with Dr Eduardo Diaz at CIB-CISC to study the molecular basis for lignin degradation in Pseudomonas putida. We have shared information about potential enzymes and genes likely to be involved in lignin degradation, and we have helped to interpret the transcriptomic data collected. We have also helped to carry out biochemical assays on enzyme candidates identified in his work. |
| Collaborator Contribution | Dr Eduardo Diaz from CIB-CISC has been a partner in the ERA-IB project. His role was to study using transcriptomics the genetic basis for lignin degradation in Pseudomonas putida, and to use this information to engineer Pseudomonas putida to produce high value chemicals from lignin breakdown. |
| Impact | Journal publications arising from the collaboration will be reported via Research Fish as & when they are published. |
| Start Year | 2015 |
| Description | Collaboration with Dr Stéphanie Baumberger (INRA) for lignin characterisation |
| Organisation | French National Institute of Agricultural Research |
| Department | INRA Versailles |
| Country | France |
| Sector | Academic/University |
| PI Contribution | We have supplied samples of lignin and biologically treated lignin to Dr Baumberger, and discussed the outputs of the lignin analysis with her. |
| Collaborator Contribution | Dr Baumberger has been a partner on the ERA-IB project. Her group has specific expertise in the characterisation of lignin via methods such as thioacidolysis, gel permeation chromatography, and quantitative OH determination via 31P NMR spectroscopy. On the ERA-IB project, her role has been to characterise the changes taking place in the lignin structure as it is degraded by Pseudomonas putida. These methods have also been applied to the study of the mechanism of action of Sphingobacterium sp. T2 manganese superoxide dismutase. |
| Impact | Journal publications will be reported via Research Fish in due course as they are published. |
| Start Year | 2015 |
| Description | Invited lecture at Lignin Gordon Research Conference, Stonehill College, Easton MA, USA, 2nd August 2022 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | I gave an invited lecture at the 2022 Lignin Gordon Conference, which is the leading international conference in the lignin valorisation field. The conference was attended by about 120 scientific researchers and industrialists in the lignin field. My presentation led to a number of questions, and one exchange of materials since the conference. |
| Year(s) Of Engagement Activity | 2022 |
| URL | https://www.grc.org/lignin-conference/2022/ |
| Description | Invited seminar at International Conference on Environmental Catalysis, 6th-9th September 2020, University of Manchester |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Postgraduate students |
| Results and Impact | I was invited as a plenary speaker to this International Conference, which due to the COVID-19 pandemic was run online rather than in-person. There was an audience of 100-150 conference delegates from many different countries, made up of postgraduate and postdoctoral researchers, research academics, and industry representatives. There were a number of questions after my seminar, which were discussed at the time. |
| Year(s) Of Engagement Activity | 2020 |
| Description | Keynote lecture at Lignocost conference, Wageningen, The Netherlands, 2nd June 2022 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | Gave an invited keynote lecture to the LignoCost network conference, attended by 120 participants from around Europe, mainly academic researchers working in the lignin field, and several industrialists from European companies. My presentation led to several conversations with academic groups, and one company from Finland, with whom I exchanged contact details. |
| Year(s) Of Engagement Activity | 2022 |
| Description | Seminar at Lignocost online meeting 26th April 2021 "Bioconversion of polymeric lignin to target bioproducts using engineered microbes" |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Professional Practitioners |
| Results and Impact | 60-70 researchers in the Lignocost network (EU COST Action) attended an online research workshop Lon lignin valorisation, where Prof Bugg presented an invited seminar. The seminar led to further questions from participants. |
| Year(s) Of Engagement Activity | 2021 |