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Predictive and Accelerated Metabolic Engineering Network

Periodic Reporting for period 2 - PAcMEN (Predictive and Accelerated Metabolic Engineering Network)

Reporting period: 2018-10-01 to 2020-09-30

The United Nations have adopted a historical agreement on reduction of carbon dioxide emissions on the 12th of December 2015. The ambition is to hold the increase in the global average temperature to well below 2°C above the pre-industrial level, which means limiting the CO2 emissions to 40 gigatones per year in 2030. This will require to double utilization of biomass and waste. A shift towards bio-based production of chemicals and fuels can significantly reduce our dependency on oil and gas and lead to a more environmentally friendly economic growth. The main challenge for industrial biotechnology is the development of novel cell factories that can efficiently convert feedstocks into the product of interest at minimum processing cost. Currently, it takes over 50 man-years and over $50 million to develop a cell factory, an investment that is difficult to finance and recover. Hence, there is a critical need for novel technologies and approaches that can change the paradigm and deliver novel cell factories faster and cheaper. For that we need researchers with interdisciplinary scientific training and with insight into the industrial biotechnology processes and business.
The European Training Network on Predictive and Accelerated Metabolic Engineering Network (PAcMEN) is training sixteen Early Stage Researchers (ESRs) in the following complementary research disciplines, which together offer an opportunity to revolutionize the development of cell factories for biotechnology:
• Systems Biology and Modeling,
• Synthetic Biology and Protein Engineering, and
• Applications for Development of New Biorefinery Processes.

The overall objective of PAcMEN research programme is to bring step-change innovation into development of cell factories for industrial biotechnology, where modeling and utilization of omics data will enable more predictive cell factory design and synthetic biology will accelerate strain construction and screening. The ultimate research aim is to cut down the cost and time of developing new cell factories.

Following 4 years of exciting ESR training, the PAcMEN ITN project is proud to conclude that it has achieved its main objectives on i) development of predictive and accelerated engineering of yeast cell factories, but also its secondary objectives on ii) completion of the training of a cadre of interdisciplinary next-generation metabolic engineers with an entrepreneurial mindset, and iii) strengthen inter-sectorial long-term collaborations.
The research conducted has covered three disciplines: metabolic modeling, synthetic biology, and metabolic engineering. The main achievements can be summarized as following.
Metabolic Modeling: the genome-scale model of yeast Saccharomyces cerevisiae was extended with thermostability and regulatory constraints, as well as transcriptional regulation information. A thermodynamic model of oleaginous yeast Yarrowia lipolytica has been constructed, including a manually curated thermodynamic model of lipid metabolism.
Synthesis Biology: The ESRs have developed i) standardized toolkits for efficient genome engineering of Yarrowia lipolytica, ii) designed, constructed and applied high-throughput and automated genotyping and phenotyping workflows for Saccharomyces cerevisiae, (iii) developed and applied protein-based biosensors and other ultra-high throughput methodologies for screening of cell factories and control of cellular metabolism, as well as iv) developed streamlined directed evolution workflows.
Metabolic Engineering: The ESRs have engineered novel yeast strains that can convert sugar into value-added products as fatty alcohols, lactones, and aromatics. Pathways of first and second generation leading have been constructed using either molecular genetics or evolutionary engineering that are either able to grow without vitamins supplementation to strains able to express new enzyme co-factors enabling expansion of their enzymatic catalogue. Moreover, regulatory control circuits have enabled growth-coupling of anabolic fitness-burdening product formation, thereby safe-guarding cell factories from product-declines under industry-mimicking conditions.
Dissemination: All ESRs have participated in international outreach activities such as conferences and workshops. Also, more than 10 ESRs have organized and participated in Open Science events for the general public, and all ESRs have proactively managed the PAcMEN social media accounts and webpage. Furthermore, currently a total of 29 peer-reviewed publication have already been published by PAcMEN ESRs and many more publications are currently in preparation. The updated list of publications can be found on the PAcMEN official website All PAcMEN papers are published with open access. The results have also been presented at over 14 international conferences, e.g. Metabolic Engienering 12, 33rd International Specialised Symposium on Yeast (ISSY33), Enzyme Engineering XXIV, and others. Moreover, the project results have been disseminated to general public through nine press releases, which have been communicated through various local and international platforms.
In terms of state-of-the-art advancement, PAcMEN has successfully advanced both the modelling, engineering and as well as the cost-effectiveness of yeast cell factory bioprocesses. This is exemplified by the high number of peer-reviewed research manuscripts already published by the ESRs, with several more in the pipeline.
PAcMEN has demonstrated a strong positive impact on the training of ESRs and on the collaborations and research advances of the involved beneficiaries and partners. During the recruitment process, many candidates expressed that PAcMEN training program was particularly attractive due to the international and inter-sectorial training that it has provided. The steering committee has also conducted anonymous satisfactory survey and also here the responses from the ESRs have been very positive. PAcMEN has allowed the academic beneficiaries to set up some unique new courses that build on the expertise of several partners, such as Winter Modeling School, Summer School on Protein Engineering and to organize a Publishing Course together with Nature Publishing Group and BioBusiness and Innovation course together with Copenhagen Business school. Such activities could be completed only thanks to the network, because of the high cost and organizational load. In summary, through international inters-sectorial training of the ESRs, new contacts and collaborations have been established between the beneficiaries and partners of the PAcMEN network. The industrial partners have benefited from hosting the ESRs, as the ESRs have contributed with ideas and product development. Some partners have or are planning to recruit the ESRs from PAcMEN, for example ESR16 has been recruited by Ginkgo Bioworks, where he has completed his 6-month secondment. Undoubtedly, PAcMEN has enabled the establishment of professional bonds which will lead to more collaborations in the future.