IN FOCUS

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Network NewsGrowing the UK Industrial Biotechnology baseThe UK Bioeconomy StrategyBBSRC - Forward Look for UK BioscienceIndustrial Biotechnology Policy – how can you get involved?BBSRC Networks in Industrial Biotechnology and Bioenergy - Phase IICBMNet Achievements - An Overview

Events & Early Career Researcher GrantsImport and Export of Small Molecules for BiocatalysisFactories for Advanced BiomanufacturingIntroduction to Industrial Biotechnology – Short CourseExperimental Techniques for Studying Proteins and Lipids in Biological MembranesCBMNet Early Career Researcher GrantsFuture Events

Business Interaction VouchersCondition-specific engineering of Pseudomonas metabolismEfflux from bacteria of feedstocks for industrial manufactureEngineering E. coli for enhanced production of antibody fragmentsPackaging proteins from bacteriaHeterologous expression of biosurfactant transporters in Debaryomyces hansenii

Proof-of-Concept FundingConsolidation, integration & critical mass building - optimizing membrane function in the Clostridial ABE processInteractions between mimetic bacterial membranesUse of membrane complexes for the production of microalgal polysaccharidesBioenergy production from biorefineries waste using super yeastsIndustrial Strategy Challenge Fund - Early Stage Feasibility Projects

Vacation ScholarshipsMarket analysis of the IBBE sector: relevant areas to Project DETOXProtein-mediated transport across hybrid lipid-block copolymer membranesDurable vesicles for stabilisation of membrane proteins in biotechnologyFunctional characterisation of a putative succinate efflux pump from Corynebacterium glutamicumExamining the potential for periplasmic volume manipulation and its influence on Sec and FT3SS based protein production

Spotlight on IndustryDr Jen Vanderhoven, FUJIFILM Diosynth BiotechnologiesDr Chris Lennon, FUJIFILM Diosynth BiotechnologiesMany thanks to our ever-supportive Management Board

Looking AheadBBSRC Phase II NIBB:

Algae-UK, BBNet, CCNet, E3B, EBNet, HVB NetworkCBMNet Themes - Our Legacy

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It is with mixed feelings that we welcome you to the final CBMNet In Focus publication. We are disappointed that EBMNet, the proposed successor to CBMNet, was not supported as one of the Phase II NIBB. EBMNet would have retained all the core strengths of CBMNet whilst forging new alliances by breaking new ground in systems metabolic engineering and would have made a major contribution to developing new partnerships and knowledge exchange for the benefit of UK Biotechnology. We wish all the Phase II NIBB well and we hope that CBMNet members, academic and industrial, can find at least one Phase II NIBB that supports their needs and interests. In the meantime, we are exploring possible ways to preserve at least some of the assets of CBMNet that were not rolled over in Phase II; hopefully there will be more news soon.

Whilst the sense of disappointment in knowing that CBMNet will close in June 2019 remains, there is also a strong sense of satisfaction in having been part of building a new community of academic researchers, biotechnology practitioners and policy makers to promote UK biotechnology. In particular, the importance of considering how membrane function impacts on the performance of any cell factory.

In this final edition of In Focus we are pleased to showcase the many ways in which CBMNet and its members have expanded knowledge and pressed the case for biotechnology in the last 12 months. The launch of the ‘National Industrial Biotechnology Strategy to 2030’ at the House of Commons in June 2018 was an opportunity to put the case for Government support for UK biotechnology directly to MPs. Hopefully, once the all-consuming focus on leaving the EU is behind us, there will be further opportunities to take this implementation plan forward with ministerial support. It has also been encouraging to see how CBMNet Business Interaction Vouchers, Proof-of-Concept funds and Vacation Scholarships have engaged academics that have not previously been involved in industrial biotechnology projects. Several exciting case studies are recounted here and some have led on to further funding to maintain these new relationships.

We hope that you find something of interest in this edition of In Focus. We thank all those who have served on the CBMNet management board for their enthusiastic support over the last 5 years and of course we thank all the members, without whom CBMNet would not have possible.

Prof Jeff Green, Director

Prof Gavin Thomas, Co-director

Dr Janet Cronshaw, Network Manager

Welcome...

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Network News

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In October 2017, a consortium of four UK NIBBs (CBMNet, BIOCATNET, P2P and C1Net) commissioned and published the report ‘Developing a UK Strategy for Industrial Biotechnology and Bioenergy’. Building on this report, an implementation plan was developed by the Industrial Biotechnology Leadership Forum (IBLF), in collaboration with CBMNet and BIOCATNET. The ‘National Industrial Biotechnology Strategy to 2030’ aims to ensure that the UK becomes a leader in the global shift towards clean growth by fostering the development of industrial biotech SMEs. It was promoted in partnership with the UK BioIndustry Association (BIA) and launched in June 2018 at the Houses of Parliament, in an event hosted by Daniel Zeichner, MP for Cambridge. The Strategy will be implemented in three phases with actions across seven key elements: external environment; access to funding and finance; infrastructure and regional footprint; trade, inward investment and commercialisation; regulations and standards; skills; communication.

Growing the UK Industrial Biotechnology BaseEnabling Technologies for a Sustainable Circular Bioeconomy: A National Industrial Biotechnology Strategy to 2030

“The Strategy describes the vision of the UK industrial biotech community, driven by the IBLF; in harnessing the world-class science we have in the UK in order to enable industrial biotech to become a mainstream part of UK industry. Through industrial biotech we can find sustainable alternatives to fossil fuels for the production of everyday chemicals, materials and energy, as well as delivering new functionalities and benefits. Industrial biotech is the only way to manufacture unique biopharmaceuticals. These help tackle serious illness such as cancer and infectious diseases. Industrial biotech can mitigate climate change through the development of greener, cleaner manufacturing processes, as well as offering opportunities for waste utilisation and new products that benefit society that cannot be made any other way.”

Steve Bagshaw, CEO of Fujifilm Diosynth Biotechnologies and Chair of the IBLF

“Industrial biotech plays a key role in societal goals like helping eliminate avoidable plastic waste which David Attenborough’s TV programme Blue Planet 2 has brought to recent public attention. The BIA is delighted that today’s strategy showcases UK entrepreneurs unleashing the innovation that will provide not only a cleaner, greener Britain but also prosperity for the next generation. This sector is key to developing the technologically and economically practical innovation needed to deliver the environmental objectives of the UK government.”

Steve Bates OBE, CEO of BIA

Useful links:NIBB Strategy: https://www.bioindustry.org/resource-listing/a-national-industrial-biotechnology-strategy-to-2030.html

NIBB report: http://cbmnetnibb.group.shef.ac.uk/industrial-biotechnology-report-launched-in-sheffield/

CBMNet: http://www.cbmnetnibb.net/ BIOCATNET: http://biocatnet.com/ BIA: https://www.bioindustry.org/ IBLF: http://www.iblfglobal.org/

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“The NIBB has brought together industry and academia with great success, with 800 companies and 3,300 academics amongst their membership. BBSRC’s Annual Report (2017) shows that £8.2m of funding had been committed as of January 2017, attracting £1.2m in private-sector investment. Building on this success by replicating across sectors will accelerate the innovation needed to solve global challenges through the bioeconomy.”

UK Bioeconomy Strategy

Useful links:UK Bioeconomy Strategy: https://www.gov.uk/government/publications/bioeconomy-strategy-2018-to-2030

BBSRC Forward Look for UK Bioscience: https://bbsrc.ukri.org/news/planning/forwardlook/

UK Industrial Strategy: https://www.gov.uk/government/topical-events/the-uks-industrial-strategy

Life Sciences Sector Deal 2 (2018): https://www.gov.uk/government/publications/life-sciences-sector-deal/life-sciences-sector-deal-2-2018

Europe’s bioeconomy strategy: https://ec.europa.eu/research/bioeconomy/index.cfm?pg=policy&lib=strategy

The UK Bioeconomy StrategyOn 5 December 2019, the UK Bioeconomy Strategy was published. The strategy is a collective approach from government, industry and the research community to transform the UK economy through the power of bioscience and biotechnology.

According to the strategy, the bioeconomy represents the “economic potential of harnessing the power of bioscience, using renewable biological resources to replace fossil resources in innovative products, processes and services”. It is estimated that the bioeconomy in the UK in 2014 contributed to £220 billion of output across the UK economy, supporting 5.2 million jobs.

The strategy has 4 main strategic goals:

1. Maximise productivity and potential from existing UK bioeconomy assets.

2. Create the right societal and market conditions to allow novel bio-based products and services to thrive.

3. Capitalise on our world class research, development and innovation base to grow the bioeconomy.

4. Deliver real, measurable benefits for the UK economy.

“The Forward Look covers a period when the research landscape is changing rapidly. There are significant opportunities for UK bioscience to drive growth in the bioeconomy, acting as a focus for private sector investments and inspiring new companies around potentially disruptive bio-based solutions in exciting areas of agri-tech, industrial biotechnology and synthetic biology to name but a few.”

Professor Melanie Welham, Executive Chair of UKRI - BBSRC.

In September 2018, BBSRC published a roadmap setting out BBSRC’s vision of the future of UK bioscience. The “Forward Look for UK Bioscience” aims to address challenges faced by future generations, such as food security, clean growth and healthy ageing, by focusing on three main themes:

1. Advancing the frontiers of bioscience discovery

2. Tackling strategic challenges

3. Building strong foundations

In Spring 2019, BBSRC is expected to publish its “Strategic Delivery Plan” which will develop the roadmap further and will guide how BBSRC invests in bioscience research, technology, infrastructure and people in the future.

BBSRC - Forward Look for UK Bioscience

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Growth Emissions Reduction Targets’. We have also made a submission to the Committee’s ‘My Science Inquiry’, an open opportunity to suggest science and technology topics for scrutiny. Our published submissions can be accessed here: https://www.parliament.uk/business/committees/committees-a-z/commons-select/science-and-technology-committee/inquiries/

For anyone interested in getting involved in science policy, you can do so via a number of organisations:

Industrial Biotechnology Policy: how can you get involved?As an organisation, we consider it a role of CBMNet to be involved in policy discussions to ensure that Industrial Biotechnology has a voice. You may remember that, in 2018, we asked for your opinion on how Industrial Biotechnology can contribute to the government’s ‘Clean Growth’ grand challenge of the Industrial Strategy. Many thanks to those of you that responded; as a result, we have submitted evidence to the House of Commons Science and Technology Committee inquiry into the ‘Technologies Needed to Meet Clean

The Biochemical Society:https://www.biochemistry.org/Sciencepolicy.aspx

The Microbiology Society:https://microbiologysociety.org/policy/get-involved.html

The Parliamentary Office of Science and Technology (POST):https://www.parliament.uk/mps-lords-and-offices/offices/bicameral/post/

The House of Commons Science & Technology Committee inquiries:https://www.parliament.uk/business/committees/committees-a-z/commons-select/science-and-technology-committee/inquiries/

BBSRC Networks in Industrial Biotechnology and Bioenergy - Phase IIMoving to a low carbon economy in the coming decades requires a shift from using fossil resources to provide power, fuel, chemicals and materials. BBSRC, with the support of EPSRC, have committed £11 million to fund six unique collaborative Networks in Industrial Biotechnology and Bioenergy (BBSRC NIBB) to support, encourage, and facilitate this essential work. The Networks build on the success of the first phase of networks, including CBMNet itself, and will run from 2019 to 2024. The new networks will provide flexible funding for Proof of Concept projects, and will be open to new members throughout their lifetime.

More details of the individulal networks can be found in “Looking Ahead”, page 37.

“These networks are testament to the success of NIBB Phase I in harnessing the UK’s world-leading industrial biotechnology. It is through industrial biotechnology that we are able to create new processes and products that are more sustainable and far better for the environment. I’m excited to see what NIBB Phase II will do to tackle the global challenges facing us all.”

Steve Bagshaw, CEO Fujifilm Diosynth Biotechnologies, Chair of IBLF

“This substantial investment into NIBB Phase II reflects the determination we have to secure a cleaner, greener, and a more sustainable future. As part of UK Research and Innovation, BBSRC continues to invest in industrial biotechnology research that underpins these networks, and will pave the way for novel and game-changing industrial processes and products.”

Professor Melanie Welham, BBSRC Executive Chair

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CBMNet Achievements - An Overview

Funding from BBSRC via CBMNetBusiness Interaction Vouchers: £207,000Proof-of-Concept Funding: £759,000Vacation Scholarships: £34,000Early Career Researcher Grants: £15,000

Industry investmentCash: £107,000In-kind: £455,000

>30 Network eventsEvent sponsorshipOpen innovation meetingsIndustrial-academic exchange grantsInternational exchange visitsPublic outreach activities

More than 1,000 membersfrom over 400 organisations

Academia: 75 %Industry: 20 %UK: 79 %International: 21 %

48 IB-related research projects24 Proof-of-Concept awards24 Business Interaction Vouchers

~£19 million funding leveraged, including...2 IB Catalyst awards2 BBSRC LINK grants3 Innovate UK awards1 ERA CoBioTechInternational partnering award15 PhD studentships

Early Career ResearchersCareers event for ECRsIB training courses21 ECRs funded to attend meetingsVacation studentships for 20 undergraduates

Publications

Public artwork

Consortia for funding applications

CBMNet project case studies: http://cbmnetnibb.group.shef.ac.uk/outputs/case-studies/

CBMNet event reports: http://cbmnetnibb.group.shef.ac.uk/members-forum/event-reports/

Publications from CBMNet projects: http://cbmnetnibb.group.shef.ac.uk/outputs/papers-published-from-cbmnet-projects/

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Funding from BBSRC via CBMNetBusiness Interaction Vouchers: £207,000Proof-of-Concept Funding: £759,000Vacation Scholarships: £34,000Early Career Researcher Grants: £15,000

Industry investmentCash: £107,000In-kind: £455,000

>30 Network eventsEvent sponsorshipOpen innovation meetingsIndustrial-academic exchange grantsInternational exchange visitsPublic outreach activities

More than 1,000 membersfrom over 400 organisations

Academia: 75 %Industry: 20 %UK: 79 %International: 21 %

48 IB-related research projects24 Proof-of-Concept awards24 Business Interaction Vouchers

~£19 million funding leveraged, including...2 IB Catalyst awards2 BBSRC LINK grants3 Innovate UK awards1 ERA CoBioTechInternational partnering award15 PhD studentships

Early Career ResearchersCareers event for ECRsIB training courses21 ECRs funded to attend meetingsVacation studentships for 20 undergraduates

Publications

Public artwork

Consortia for funding applications

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Events & Early Career Researcher Grants

20 Early Career Researchers8 institutions

£17,000 from CBMNet£3,000 in-kind from industry

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Import and Export of Small Molecules for BiocatalysisEdinburgh | September 2017

This joint CBMNet and BIOCATNET event brought together over sixty scientists with diverse interests in the import and export of small molecules. The event was well attended by academics and included high quality talks from Phillip Stansfeld (University of Oxford), Kostas Beis (Imperial College London), Gary Black (Northumbria University) and Beppe Battaglia (UCL). Complementing the academic input were contributions from industrial scientists at Ingenza, GlaxoSmithKline and IBioIC, who are interested in improving the robustness of whole-cell biocatalysis to overcome restrictions on product titres, separability, sustainability and process economics.

“The high quality of science was remarkable, and the fact that you had people facing the same problems from (very) different perspectives gave new ideas and insight on how to address my own limitations.”

“The best part of the event was the ability to communicate openly and honestly regarding current challenges faced by both academics and industry. For instance, it was beneficial to discuss some discrepancies between the focus of academic research in contrast with the current limitations in industrial settings.”

“I really enjoy the relevant NIBB events as they keep ideas fresh and my science up to date and to get to know experts in the field.”

This event brought together over 130 scientists with diverse interests in microbial chassis engineering, alongside industrial scientists who are interested in improving the robustness of whole-cell biocatalysts. Key themes for the event addressed six major challenges in chassis design:

• Reverse-engineering of microbial chassis, where the manufacturing process is designed first, and the biocatalyst is developed to match the process constraints

• Tolerance to toxic products and substrates, to overcome current restrictions on product titres

• Improved strain stability for continuous manufacturing, to achieve the productivity critical for bulk bio-based chemicals manufacturing

• Improved atom efficiency, to enable efficient conversion of substrate to product without loss of carbon, by diverting fluxes away from cell growth towards product and minimizing by-product formation

• Improved transport processes, to ensure efficient substrate uptake and product efflux

• Innovative high throughput tools for systems metabolic engineering, including CRISPR-Cas9, integrase technology, directed evolution, high-throughput screening and selection.

Academic speakers included Gavin Thomas (University of York), Colin Harwood (Newcastle University), Dave Kelly (University of Sheffield), James Allen (UCL), Mike Lynch (Duke University), Claudio Angione (Teesside University), Gill Stephens & Jon McKechnie (University of Nottingham), John Heap (Imperial College London) and Susan Molyneux-Hodgson (University of Exeter). Flash presentations were selected from researchers at the Universities of Sheffield, Leeds, Nottingham, Manchester and Aberystwyth and from the International Centre for Genetic Engineering and Biotechnology in New Delhi.

As always, industrial contributions to CBMNet meetings are highly valued and contributions were welcomed from VideraBio, Oxford Biotrans, Green Biologics Ltd., Syngulon, CHAIN Biotechnology Ltd., Ingenza Ltd., Biocatalysts Ltd., Isobionics BV, Calysta UK, Matis and NNFCC.

Factories for Advanced BiomanufacturingSheffield | December 2017

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Introduction to Industrial Biotechnology – Short CourseMiddlesbrough | March 2018

Building on the success of the first training course in November 2016, CBMNet organised a second 3-day course in Industrial Biotechnology and Bioenergy. The course provided 18 delegates with a foundation in the understanding of biological membranes and the role they play in Industrial Biotechnology, via a series of lectures from key academics and industrial speakers. Delegates had the opportunity to present their own research and also participated in problem solving case studies and a journal club.

Included in the course was a tour of the Wilton Centre at The Centre for Process Innovation as well as a tour and laboratory practicals at Fujifilm Diosynth Biotechnologies.

“The CBMNet course provided me with a refresher in stabilised biochemical techniques and an insight into new methods. It also offered the opportunity to build connections with academics and industry members working on complementary topics. A thoroughly worthwhile course!”

“I found this event interesting and stimulating. I came away with new ideas for research and new contacts. I would certainly recommend this event to anyone new to, or interested in developing links and research industrial biotechnology.”

“The short course was really informative and I was intrigued by the vast opportunities to exploit the huge potential that advances in Industrial Biotechnology offers.”

“The 3 days was an excellent way to get up-to-date information on current membrane biology techniques from a mixture of academic and industrial scientists. The site tours and practical demonstrations showed how the scale-up of processes from the lab bench to demonstration/pilot scale should be done.”

Experimental Techniques for Studying Proteins and Lipids in Biological MembranesBirmingham | July 2018

Many thanks to the Biochemical Society for organising this CBMNet-sponsored meeting. Thanks must also go to the programme coordinators, Kostas Beis and Alan Goddard, who did a great job developing a stimulating agenda of talks. Over the two days, we heard from Alice Rothnie and Irundika Dias (Aston University), Manuela Mura (University of Lincoln), Gavin Thomas (University of York), Kostas Beis (Imperial College London), Saskia Bakker (University of Warwick Advanced Bioimaging Research Technology Platform) and Argyris Politis (King’s College London). The training event also offered practical sessions on the preparation and application of SMALPs, expertly led by Alice Rothnie (Aston University). It was great to have Liz Jenkinson of Green Biologics at one of these sessions to contribute to a discussion on the challenges and advantages of working in or with industry.

A follow-up event is being organised for April 2019 (see future events or the CBMNet Events webpage)

“This meeting was a thorough and engaging introduction to a wide variety of techniques. The speakers gave in-depth advice on the use of different techniques. It was also great to get some practical experience mixed in with the lectures!”

Useful links:For future events that may be of interest to you, see: http://cbmnetnibb.group.shef.ac.uk/events/

For reports on past CBMNet events, see: http://cbmnetnibb.group.shef.ac.uk/members-forum/event-reports/

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CBMNet Early Career Researcher GrantsIn February 2018, CBMNet sponsored three of our members to attend “The artificial cell: biology-inspired compartmentalisation of chemical function”. This Royal Society Theo

Murphy meeting was held at the Kavli Royal Society Centre, Chicheley Hall in Buckinghamshire and was organised by Paul A. Beales (University of Leeds), Barbara Ciani (University of Sheffield), and Stephen Mann (University of Bristol). CBMNet members, Rashmi Seneviratne and Andrew Booth (University of Leeds) and Daniel Mitchell (University of Sheffield) all presented posters at the event and you can see their full reports on our website:http://cbmnetnibb.group.shef.ac.uk/outputs/case-studies/early-career-researcher-grants/the-artificial-cell-biology-inspired-compartmentalisation-of-chemical-function/

The Royal Society also published a theme issue of Interface Focus, based on the outputs from this meeting:https://royalsocietypublishing.org/doi/full/10.1098/rsfs.2018.0046

In April 2018, CBMNet sponsored two of our members to attend the European Federation of Biotechnology (EFB) Microbial Stress Meeting: From Systems to Molecules and Back in Kinsale, County Cork, Ireland. Arthur Neuberger (University of Cambridge) and Aidan Taylor (University of Sheffield) both gave talks at the meeting on various aspects of “Stress at the Systems and Structural Level”. Both reported a successful, varied and interesting meeting and success in making potentially valuable new contacts. See our website for the full event report:http://cbmnetnibb.group.shef.ac.uk/outputs/case-studies/early-career-researcher-grants/efb-microbial-stress/

“CBMNet’s generous funding allowed me to attend a meeting where I was exposed to a myriad of ideas, beyond my immediate field, which will help shape my future career.”

Future Events

30 April – 2 May 2019 | Sheffield, UK

CBMNet’s final event, “Industrial Biotechnology at the Cell Membrane” will focus on four main themes:

• Uptake

• Efflux & Membrane Technologies

• Membrane Engineering/Chemicals Toxicity

• Protein Secretion

We have some great speakers confirmed, including the CBMNet directors Jeff Green and Gavin Thomas. For more details, and to register, see the CBMNet events website:http://cbmnetnibb.group.shef.ac.uk/registration-open-industrial-biotechnology-at-the-cell-membrane-30-april-2-may-2019/

A Membrane Protein Training Event is planned to follow on from the highly successful “Experimental Techniques for Studying Proteins and Lipids in Biological Membranes”. CBMNet is sponsoring a second Biochemical Society training event to be held at the University of Leeds:

2nd UK Workshop on Membrane proteins: solubilisation and biophysical characterisation

3-5 April 2019 | University of Leeds, UK

For more details, and to register, see:https://www.biochemistry.org/Events/tabid/379/MeetingNo/TD027/view/Conference/Default.aspx

Other Events:8-11 April 2019: Microbiology Society Annual Conference 2019 (Belfast, UK)

26 April 2019: SMALP 2019 (Utrecht, Netherlands)

22-28 June 2019: Gordon Research Seminar and Conference – Mechanisms of Membrane Transport (New London, NH, USA)

28-31 August 2019: 85th Harden Conference: Dynamic Membrane Complexes: Respiration and Transport (Bonn, Germany)

9-10 December 2019: Synthetic Biology UK 2019 (Warwick, UK)

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Business Interaction Vouchers

24 projects22 academics

13 institutions17 companies

£207,000 from CBMNet£20,000 cash + £204,000 in-kind from industry

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The ChallengeMicro-organisms have a high capacity of synthesizing a wide range of surface-active compounds, generally called biosurfactants. Rhamnolipids are a common biosurfactant and due to their non-toxic nature, excellent biodegradability, high surface/interfacial activity, high thermal/chemical stability, production from renewable resources and the ability to form micro-emulsions, rhamnolipids can be used as emulsifiers and stabilizers in a great number of sectors. Enhanced production of rhamnolipids is key for industrial applications, ranging from cosmetics to bioremediation of organic and heavy metal contaminated environments.

Rhamnolipids are predominantly produced by the microbe Pseudomonas aeruginosa. However, Pseudomonas putida is a model organism with greater metabolic versatility and potential for industrial applications. Computational methods for metabolic engineering are able to model and optimise such biological models, leading to the improvement of a given biotechnological pipeline.

The ResearchDr Angione is a Senior Lecturer at Teesside University. The research in his laboratory focuses mainly on computational biology and modelling for biomedical applications.

TeeGene Biotech Ltd is a spin-out venture from Teesside University, which is pioneering the use of biosurfactants in a range of household, environmental, cosmetic and biomedical applications.

Dr Angione applied for a CBMNet Business Interaction Voucher with TeeGene Biotech Ltd. to develop a collaborative partnership and produce preliminary data that could be built on in future funded projects. The project aimed to investigate the metabolic capabilities of P. putida for rhamnolipid

Condition-specific engineering of Pseudomonas metabolismClaudio Angione, Teesside University & TeeGene Biotech Ltd.

biosynthesis using metabolic and biosynthetic engineering approaches. The analysis would be based on computational modelling and metabolic engineering.

The ResultThe engineered genome-scale model of P. putida built during this BIV project can already be used to predict and maximise rhamnolipid production and transport through the cell membrane. The P. putida model was optimised to maximise the production and export of biomass and rhamnolipids. By introducing the non-native genes RhlA and RhlB (from P. aeruginosa), P. putida can produce rhamnolipids. The project found the majority of rhamnolipid production originated from the rhamnose pathway rather than from the FA pathway.

To establish the best growth condition for optimal rhamnolipid synthesis, the project investigated alternative carbon sources separately: fructose, sucrose, glycerol, benzoate and myristic acid. Findings report that metabolism of myristic acid (C-14), followed by fructose and sucrose/glucose, provided the best condition for optimal rhamnolipid synthesis.

The FutureThe promising results found during this project have strengthened the collaboration between TeeGene and the Computational Biology group at Teesside University. Results will be extended with poly-omics modelling and validated in an upcoming project for which we are seeking funding as a Proof of Concept.

The resulting pipeline will finally be used to build a genome-scale model of Pseudomonas teessidea, first isolated and characterised by TeeGene, for which a model is unavailable. The resulting project will elucidate the metabolic engineering steps for overproduction of rhamnolipids and their transport out of the cell membrane.

“The funding awarded by CBMNet has kick-started a fruitful collaboration with TeeGene Biotech. This collaboration has been extremely successful and has already generated a number of additional interdisciplinary projects, blending computational and mathematical tools with industrial biotechnology expertise.”

Dr Claudio Angione, Teesside University

“This BIV work with Teesside University uncovering some of the challenges we face over the years to identify and optimise rhamnolipid production. This work integrates TeeGene’s core technologies and business aspirations with biotechnology and bioprocessing expertise at Teesside to create a central enabling technology for biomanufacturing of biosurfactants.”

Dr Pattanathu Rahman, TeeGene Biotech Limited

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Efflux from bacteria of feedstocks for industrial manufacturePeter Henderson, University of Leeds & Lucite International

The ChallengeA significant global challenge is to develop bioprocesses for the sustained manufacture of important commodity chemicals, whilst reducing reliance on fossil fuels. One such strategy uses microbial cell factories that are capable of making key chemicals from sugars. The development of these cost-competitive, commercially viable bioprocesses presents a significant challenge to the chemicals industry.

As a substitute for the synthetic production of commodity chemicals, Lucite International is carrying out research to utilise bacteria to host a variety of metabolic pathways to produce the required end-products from sugars. However, the end-products can be toxic to the host cells, so the challenge is to find and develop protein-mediated efflux systems that will actively export the end-product from the cell and reduce its detrimental effect on the bacteria. To achieve this requires the development of existing and novel assays to measure the efflux of the desired compound out of the microbial cell factory.

The ResearchPeter Henderson is Professor of Biochemistry and Molecular Biology at the University of Leeds. The research in his laboratory focuses mainly on understanding the molecular basis of membrane transport processes, and particularly the roles of energised efflux of toxins in resistance of micro-organisms to antibiotics.

At a CBMNet meeting, Professor Henderson gave a presentation on bacterial efflux systems, which attracted the interest of Dr Graham Eastham and Dr David Johnson from Lucite International. As an outcome of this, an applied 3-year PhD studentship was agreed between Lucite and the University of Leeds.

Prof Henderson and Dr Eastham also went on to apply for a CBMNet Business Interaction Voucher (BIV) and CBMNet Vacation Scholarship with Lucite International to develop a strategy for identifying bacterial efflux proteins that could be used to drive desired, but toxic, end-products of engineered synthetic pathways out of the host cells.

The ResultJacob Edgerton, a graduate with First Class Honours in Microbiology from Cardiff University, was selected to undertake a PhD degree supervised jointly by the University of Leeds and by Lucite.

With support from the CBMNet BIV, an information analyst generated a substantial list of promising efflux systems from a variety of bacterial species. Screening of these is ongoing. A second outcome was the identification by Dr Eastham and Prof Henderson of routes for the chemical incorporation of 14C, and possibly 3H, into molecules of relevance to the project for the development of direct assays of transport activity.

In parallel, Prof Henderson, Dr Eastham and an undergraduate student, Gabriel Hoppen, supported by a CBMNet Vacation Scholarship, explored indirect methods to measure antibiotic efflux, with a view to their application to this project.

The FutureProf Henderson and Lucite will, in the near future, be cloning and characterising a range of the efflux systems identified, which is very promising for the aims of this project. In parallel, they are exploring indirect assays for measuring efflux that involve fluorescence and/or absorbance using Acriflavine, Nile Red, Ethidium and proprietary compounds and direct measurements of transport using synthesised 14C-labelled compound(s).

In addition, through the connection of Lucite with wider networks of BBSRC- and CBMNet-funded projects, Prof Henderson and Dr Eastham are now working closely with groups at the University of Nottingham and Ingenza.

“CBMNet has proven to be extremely valuable as a way of connecting with experts in the field of microbial membrane science. Furthermore the pump priming of these interactions through BIVs, PoC and vacation studentships allows potential collaborators a low cost look-see at each other. For Lucite this has been valuable and has led to longer term more established programs which would have been difficult to justify without the aforementioned mechanisms”

Dr Graham Eastham, Lucite International

“This project was conceived at the inaugural open meeting of CBMNet in Sheffield, where I first met Lucite International. Funding for a studentship was then attracted from Lucite International and the University of Leeds, with vital initiating elements supported by the BIV and Vacation Scholarship from CBMNet. We hope that CBMNet will continue and promote similar unexpected and successful liaisons between academia and industry.”

Professor Peter Henderson, University of Leeds

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Engineering E. coli for enhanced production of antibody fragmentsMark Shepherd, University of Kent & FUJIFILM Diosynth Biotechnologies

The ChallengeE. coli-based production of therapeutic proteins often requires that the products are exported to the more oxidising environment of the periplasm in order to acquire the disulphide bonds that are necessary for proper function. One example of this is the production of antibody fragments which have wide ranging applications in diagnostics, human therapeutics and as fundamental research tools.

A major problem with industrial production of antibody fragments arises when disulphide bonds are not incorporated properly, leading to diminished quality and value of the end product. To address this issue, this project aimed to exploit the E. coli ABC transporter CydDC, which exports low molecular weight thiols such as cysteine and glutathione to the E. coli periplasm where they can participate in disulphide exchange and impact upon the formation of disulphide bonds.

The ResearchDr Mark Shepherd is a Senior Lecturer in Microbial Biochemistry at the University of Kent. The research in his laboratory focuses mainly on bacterial stress tolerance, respiratory metabolism, haem biosynthesis, disulphide folding, and biofuel production.

FUJIFILM Diosynth Biotechnologies (FDB) are a world leading cGMP Contract Development and Manufacturing

Organization (CDMO) supporting the biopharmaceutical industry with the development and production of their therapeutic candidates.

Dr Shepherd applied for a CBMNet Business Interaction Voucher with FDB. Their project aimed to measure the quality of antibody fragments produced in strains of E. coli with varied CydDC levels.

The ResultThe main focus of this work became the development of a quantitative technique to measure disulphide folding in antibody Fab fragments. A state-of-the-art Waters Synapt G2-Si electrospray mass spectrometer was used to directly measure the incorporation of disulphide bonds into D1.3 Fab, a model antibody fragment used routinely by the industrial partner.

D1.3 tryptic-digest peptide fragments were analysed from E. coli strains expressing various levels of the CydDC transporter. The extent to which D1.3 was disulphide bonded was quantified for each strain, and the data revealed that loss of CydDC expression elicited a 25 % increase in antibody quality (in terms of disulphide incorporation).

The FutureDiminished CydDC expression was shown to enhance disulphide incorporation in D1.3 Fab, hence improving protein quality. This approach will be extended to other disulphide-containing protein targets of biotechnological importance in future.

The collaboration and data that has come out of this project will lead to further collaboration between Dr Shepherd and FDB. This collaboration has recently been supported by funding from the ‘Metals in Biology’ NIBB, and the project partners are currently planning antibody experiments using the approaches described herein.

“This could be of major importance to Diosynth if this approach could be extended to other Fab fragments that are expressed in high abundance when CydDC is co-expressed”

Dr Chris Lennon, Fujifilm Diosynth Biotechnologies

“This work is of value to our research as it resulted in method development in terms of proteomics approaches to measure disulphide incorporation and quantification of low molecular weight thiols. These approaches will certainly be exploited for other projects involving disulphide folding”

Dr Mark Shepherd, University of Kent

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Packaging proteins from bacteriaDan Mulvihill, University of Kent & FUJIFILM Diosynth Biotechnologies

The ChallengeThe isolation of recombinant biopharmaceuticals from large scale fermentation cultures is a major challenge to industry. The ability to target secretion of recombinant proteins into media is attractive as it improves efficiency and reduces costs. Currently there are no methods for transporting recombinant proteins across the inner and outer bacterial membrane, or for generating vesicles to facilitate the subsequent isolation and purification of the biopharmaceutical from the culture media.

The ResearchDr Dan Mulvihill is a Reader in Cell and Molecular Biology at the University of Kent. In the course of research into the dynamic re-organisation of cellular components within living cells, his group identified a protein which, when expressed in the Gram-negative bacteria, E. coli, drives the formation of extracellular vesicles, or exosomes. This provided an exciting potential opportunity for delivering recombinant proteins from the bacteria, into the culture media, to allow simple protein purification, without needing to harvest the cell culture.

The ResultWorking in collaboration with Chris Lennon, at FUJIFILM Diosynth Biotechnologies, Dan’s group generated a series of constructs to test whether this technology could be used to target proteins of interest into the vesicles in both small-scale laboratory and large scale industrial fermentation cultures of E. coli. Using both biochemical and structured illumination microscopy super-resolution imaging techniques, they showed that they were able to deliver the target protein into the cellular vesicles, which could be harvested from the media of active batch scale fermentation cultures.

The FutureDr Mulvihill has generated exciting proof-of-concept data, that show it is possible to generate synthetic vesicles into which proteins of interest can be specifically targeted, to facilitate their subsequent purification. The study has the potential to be further developed to generate membrane vesicles for drug delivery, vaccination and biopharmaceutical applications.

Dr Mulvihill and co-investigator Jennifer Hiscock, also from the University of Kent, were subsequently successful in their application for a BBSRC ‘Stand-alone’ LINK grant. This will fund their ongoing collaboration with FUJIFILM Diosynth Biotechnologies, building on the research started in this CBMNet Business Interaction Voucher.

“The CBMNet network and subsequent BIV funding has enabled me to increase the impact and potential of a discovery made in the course of a BBSRC funded research project. They have led to the development of an exciting new industrial collaboration and have allowed us to acquire proof of concept data validating our system for producing recombinant vesicles for diverse biotechnology applications, and provided the basis for further major research funding applications and future outputs.”

Dr Dan Mulvihill, University of Kent

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Heterologous expression of biosurfactant transporters in Debaryomyces hanseniiEwald Hettema, University of Sheffield & Croda

“Due to their tractable nature, model micro-organisms are attractive candidates for industrial processes. However, limitations in their inherent biological capability lead to challenges in production & commercialisation. Organisms such as D. hansenii perform industrially attractive biochemistry but require projects such as this to develop the tools to manipulate them. I look forward to continued collaboration with Dr Hettema and Dr Gilmour.”

Dr Jeremy Bartosiak-Jentys, R&D Manager, Biotechnology, Croda

The ChallengeSurfactants are ubiquitous performance ingredients that allow formulators to create the products we all use. Although surfactants are often made from petrochemical sources, there is increasing global demand to shift to production using sustainable, biological resources and technologies. Some microbes, including certain yeast, can produce and secrete biosurfactants, often to high levels. However, manipulating synthesis to produce specific structures has proved challenging.

In order to produce biosurfactants in an industrial setting, a robust and tractable production host is required. One potential organism is the halotolerant, non-pathogenic yeast Debaryomyces hansenii, which naturally accumulates high levels of lipids, that can provide the hydrophobic portion of the surfactant molecule.

The ResearchDr Ewald Hettema is a Reader at the University of Sheffield. The research in his laboratory focuses mainly on the mechanisms underlying peroxisome biogenesis. Dr Hettema and his colleague Dr Jim Gilmour (also from the University of Sheffield), have a long-standing collaboration to investigate the physiology and molecular biology of D. hansenii.

Croda is a British company that creates, makes and sells speciality chemicals that deliver real benefits to a range of diverse products across personal, health, crop and home care markets.

Dr Hettema and Dr Gilmour applied for a CBMNet Business Interaction Voucher with Croda. Their project aimed to

engineer D. hansenii to express known fungal transporters that may play a role in improving the biosurfactant production capability of the organism.

The ResultTools for protein expression and/or tagging are limited in D. hansenii. This project successfully generated a number of novel resources that will be useful for the future development of D. hansenii as an industrial organism. In the first instance, a stable shuttle plasmid, capable of replication in both E. coli and D. hansenii, was engineered. Essential plasmid elements have also been identified and cloned, including: (1) a strong, constitutive promoter; (2) an auxotrophic marker; (3) fluorescent tags; (4) immunolabelling tags.

Since D. hansenii translates the CTG codon into both leucine and serine, instead of just leucine, all of these genes were engineered accordingly. In addition, an ORF encoding a biosurfactant transporter was generated, again with appropriate codons for expression.

The FutureIn the immediate future, important next steps will involve proving that it is possible to express a heterologous transporter in D. hansenii. The tools needed to do this are now in place and Dr Hettema plans to expand the resources at their disposal by, e.g., evaluating different promoters.

In the longer term, the tools generated in this project will be of general use to all researchers using CTG clade yeast as model organisms and should allow its development as an industrial biotechnology organism.

Croda are particularly interested in engineering an alternative robust production host for the biosynthesis of biosurfactants. To this end, Dr Hettema and Croda are actively continuing their collaboration and have successfully applied for Impact, Innovation & Knowledge Exchange funding from the University of Sheffield. They plan to further explore future research directions and how to fund their collaboration in the long-term.

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Proof-of-Concept Funding

24 projects34 academics

19 institutions19 companies

£759,000 from CBMNet£86,000 cash + £247,000 in-kind from industry

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Consolidation, integration and critical mass building - optimizing membrane function in the Clostridial ABE processAlan Goddard, Aston University & Green Biologics

The ChallengeSolventogenic Clostridia are used by Green Biologics in the commercial production of n-butanol by fermentation of sugar-containing feedstocks via the acetone-butanol-ethanol (ABE) process. n-butanol is an important commodity chemical and its biological production decreases greenhouse gas emissions compared to petrochemical routes. The cost of n-butanol extraction during fermentation is high, but could be significantly decreased if higher concentrations could be achieved during fermentation. However, n-butanol over ~2% is toxic to Clostridia and even low concentrations slow metabolism.

Previous research (supported by CBMNet funding) has demonstrated that this toxicity is at least in part due to damage to the Clostridial cell membrane. However, the Clostridial membrane remains poorly understood and it is necessary to better characterise this membrane to understand how to mitigate toxicity.

The ResearchDr Alan Goddard is a Lecturer at Aston University. The research in his laboratory focuses mainly on the lipid membrane that surrounds biological cells and the integral proteins residing within this. Green Biologics makes renewable specialty chemicals and formulated products that are used by manufacturers in the fast moving consumer goods and industrial product sectors to make high performing products from renewable sources. Dr Goddard and collaborators Prof Gavin Thomas (University of York), Dr Rob Fagan (University of Sheffield) and Dr Peter Chivers (Durham University) applied for a Proof-of-Concept award with Green Biologics. This project aimed to gain further insight into the effect of n-butanol on membrane structure and function to ultimately generate resistant Clostridial strains.

The ResultSignificant progress has been made towards the aims of this project: generation of a Clostridial knockout library to screen for strains that have enhanced resistance to n-butanol; characterisation of carbohydrate transporters within the cell membrane to improve utilisation of different feedstocks; and investigation of the effects of n-butanol on metal homeostasis. A number of researchers from the host laboratories have gained experience in new techniques as well as benefiting from undertaking inter-sectoral collaborations. This project has developed a number of interactions between the partners which will be explored both informally and through future funding applications.

The FutureNewly isolated mutant strains are available that can be characterised to assess their ability to use different feedstocks more efficiently. Preliminary characterisation of membrane protein stability is ongoing using SMALP and/or bicelles with a view to examining the effect of lipid composition on membrane protein stability in response to n-butanol. All of the collaborations will continue through informal routes using existing researchers in the PI’s laboratories. Dr Goddard and Green Biologics are currently applying for further funding to continue their collaboration and, upon completion of the outstanding areas of this collaboration, will explore the possibility of further funding through routes such as BBSRC and Innovate UK.

“This project was a great opportunity to bring together various CBMNet-funded projects to form a multidisciplinary collaboration and generate some exciting data to be used for future funding applications”

Dr Alan Goddard, Aston University

“This project has enabled us to consolidate partnerships with academics across various specialisms. The results are applicable to overcoming challenges in an industrial setting in particular tolerance of microbes to inhibitors. CBMNet funding opportunities and networking events have been instrumental in driving this research forwards”

Dr Liz Jenkinson, Green Biologics

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Interactions between mimetic bacterial membranesWuge Briscoe, University of Bristol & Procter & Gamble

The ChallengeThe World Health Organization and the Centre for Disease Prevention and Control have made drug-resistant bacteria a priority research area. One of the main challenges facing development of new antimicrobial agents (AMAs) is the lack of accurate mimetic bacteria membrane models, posing a barrier to progression due to the lack of understanding of the mode of action of AMA compounds. This affects our ability to design effective AMAs. Physical disruptions of the bacterial membranes are a direct and effective bactericidal route, and thus one of the important prerequisites for the development of next generation AMAs is to understand how the pathogen coats will interact with the AMAs. Thus, there is an urgent need for development of mimetic membranes that capture the structural and compositional sophistication of bacterial outer membranes.

The ResearchDr Wuge Briscoe is a Reader in Physical Chemistry at the University of Bristol. The research in his laboratory focuses on the surface forces mediated by surfactants, polymers and nanofluids, and on fundamental aspects of biolubrication and nanotoxicity. Proctor and Gamble (P&G) make quality personal care products that improve people’s lives.

Building on previous collaborations supported by CBMNet-funding, Dr Briscoe applied for a CBMNet Proof-of-Concept award with Dr Eric Robles at P&G Newcastle Innovation Centre, an R&D facility of the world’s largest consumer products manufacturer. The central aims of this project were to: (1) design, fabricate and characterise the inner leaflet of bacterial membranes using solid-supported asymmetric lipid membranes; (2) study the fusion of these mimetic membranes using the Surface Force Apparatus (SFA).

The ResultIn order to optimise the design of asymmetric lipid membranes, Π-A isotherms were obtained of varying membrane systems. These were then characterised using BAM imaging and by X-ray reflectivity (XRR). The results showed that incorporation of cardiolipin in the lipid mixtures had effects on lipid tail orientation and created much smaller and more stable liposomes.

Initial SFA measurements were undertaken, starting out with DOPE monolayers, and LPS-Ra monolayer adsorbed onto DOPE. The initial SFA results showed that DOPE dry monolayers are a separation distance of about 3-4 nm. Once LPS-Ra had been deposited, using the Langmuir-Schaefer approach, the thickness of the supported lipid bilayers is around 4.5 nm (in water).

The FutureIn the future, membranes will be further characterised using XRR to understand how lipid bilayers differ depending on the lipid composition.

The initial SFA studies will be built upon by varying the composition of the initial monolayer. Currently in progress is a detailed AFM study varying the temperature, CaCl2 concentration, and incubation time of LPS-Ra on different membrane systems in order to study the stability and adsorption of LPS-Ra in more detail. Studies are also underway that will compare the deposition of LPS-Ra on different membrane monolayers using XRR.

The project has opened up opportunities for further investigations relevant to fundamental research as well as product formulations.

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Use of membrane complexes for the production of microalgal polysaccharidesEgbert Hoiczyk, University of Sheffield & GlycoMar

The ChallengeThe utilisation of microalgae for production of high value chemicals has seen major advances in the last decade. A key limitation is the yield of target products, which can restrict their commercial viability. This is particularly the case with exopolysaccharides (EPS), which are produced by many microalgae and represent a large biochemically diverse resource. Although methods for the production of EPS exist, increased yield would greatly improve its industrial potential.

In Prasinococcus capsulatus, EPS appears to be synthesised in the Golgi apparatus and secreted through an adjacent transmembrane pore complex called the decapore. In order to increase EPS production, we need to better understand its transmembrane synthesis and secretion, including the precise role of the decapore in EPS synthesis, maturation, and secretion.

The ResearchDr Hoiczyk is a senior lecturer at the University of Sheffield. The research in his laboratory uses high-resolution light and electron microscopy to study the structure, dynamics and functions of important bacterial subcellular complexes.

GlycoMar is a biotechnology company, discovering and developing products for the healthcare and personal care markets.

Dr Hoiczyk applied for a CBMNet Proof-of-Concept award with GlycoMar. Their collaboration aimed to maximise future production of microalgal EPS, a product patented for use in healthcare and skin care, by better understanding its synthesis and secretion. Specifically, the goal of this project was to isolate and purify the decapore complex and to identify its protein constituents.

The ResultA protocol for using cryo-electron microscopy to re-investigate the ultrastructure of the decapore was successfully developed, revealing further structural details of EPS secretion. Surprisingly, this data has fundamentally changed the concept of EPS secretion in P. capsulatus. Based on these observations, it became clear that the decapore is not involved in this process at all. Instead, it now appears plausible that the EPS is initially secreted at the cell surface into the space between cell wall and cytoplasmic membrane.

This newly gained insight into EPS secretion in P. capsulatus has benefited GlycoMar by identifying the key process involved in the synthesis and “secretion” of EPS, thereby preventing futile efforts studying the decapore complex.

The FutureGlycoMar are now in a position to optimize polymer production by systematically testing different growth regimens and their effects on the amount of formed material as well as the kinetics of its release. Mathematical modelling and quantitative experiments will be used to determine the optimal growth conditions and time point for harvesting the algae.

Following on from this collaboration, Dr Hoiczyk and GlycoMar will explore future avenues to receive funding for a continuation of their partnership. Dissemination of their data is planned in forthcoming publications.

“This collaboration was eye opening for me with respect of industry’s research needs (challenges?)”

Dr Egbert Hoiczyk, University of Sheffield

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Bioenergy production from biorefineries waste using super yeastsVincent Postis, Leeds Beckett University & Longueteau Distillery

The ChallengeBioethanol refineries generate large amounts of liquid waste called vinasse (10 - 15 L of vinasse per litre of distilled ethanol). However, current methods of disposal of vinasse can lead to major environmental issues.

The main component of vinasse is glycerol, a potentially inexpensive alternative substrate to glucose for fermentation processes. Identifying yeast strains capable of using glycerol-based products, such as vinasse, would reduce the environmental impact of bioethanol sugarcane refineries, reduce the toxicity of this by-product and be cost-effective for fermentation industries.

The challenge is to identify yeast strains that combine enhanced ability to grow in glycerol-based products with the ability to produce added-value products, such as free fatty acids/neutral lipids.

The ResearchDr Vincent Postis is a Reader at Leeds Beckett University. The research in his laboratory focuses on a wide range of transporters of biomedical interest with a particular emphasis on the structure/function relationship of nucleoside transporters and biofermentation processes.

Dr Postis and his collaborators Dr Celia Ferreira (University of Leeds) and Dr Carine De Marcos Lousa (Leeds Beckett University) applied for a CBMNet Proof-of-Concept award.

The project aimed to identify yeast strains adapted for growth on distillery waste and capable of converting these glycerol-based feedstocks into added-value products.

The ResultThree yeast strains were subjected to random mutagenesis to identify strains better adapted for growth on medium mimicking vinasse: Saccharomyces cerevisiae (grows poorly on glycerol but is the preferred organism used in industrial biotechnology) and two strains of Yarrowia lipolytica (efficiently metabolises glycerol with high yields of neutral lipids).

Random mutagenesis of Y. Lipolytica identified three strains that grow better on synthetic medium mimicking vinasse; preliminary data suggests they also have enhanced growth on vinasse. Random mutagenesis of S. cerevisiae did not identify potential strains of interest, but progress has been made towards directed metabolic engineering of S. cerevisiae.

The microbiome of mature vinasse (sourced from Longueteau Distillery) was also investigated as a potential source of novel strains, with one yeast strain being isolated. Identification of this strain is currently underway.

The FutureY. Lipolytica strains of interest are being sequenced in order to identify the relevant mutations. The Whole Genome Sequencing approach being taken has resulted in a new collaboration between Dr Postis and other researchers. Engineering S. cerevisiae to optimise growth on vinasse and production of fatty acids is in progress.

This collaboration has formed the basis of an ongoing MRes project, building on these initial results. Dr Postis and his collaborators have applied for follow-up funding to use these strains for fermentation of agricultural wastes in Cote d’Ivoire as a part of the GCRF for Industrial Biotechnology and Bioenergy in the developing world.

The prospect of being able to reuse the vinasse for biofuel production was of great interest to industrial collaborators, paving the way for future collaboration.

“Biofermentation processes have been at the heart of human life since Egyptian times with wine and bread. Our understanding of molecular mechanism of fermentation is now the basis for the development of sustainable energy”

Dr Vincent Postis, Leeds Beckett University

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Industrial Strategy Challenge FundIndustrial Biotechnology Catalyst Early Stage Feasibility Projects

In April 2018, BBSRC awarded CBMNet over £230,000 from Wave 1 of the Industrial Strategy Challenge Fund (ISCF) to support Early Stage Feasibility Projects, within the strategic scope of the Industrial Biotechnology (IB) Catalyst. The funds were distributed to seven partnerships between academic researchers and industrial partners, who undertook Proof-of-Concept projects on a variety of topics related to the CBMNet remit of crossing biological membranes. The seven funded projects are:

Metabolic engineering of Yarrowia lipolytica for co-production of xylitol and lipids fuels using industrial wasteGary Leeke (Cranfield University) & SERE-Tech Innovation Ltd

In silico genome-wide modelling and metabolic engineering of Pseudomonas strains for improved rhamnolipid synthesisClaudio Angione (Teesside University) & TeeGene Biotech Limited

Detoxification of cyclic monoterpene alcohols by selective esterificationDavid Leak (University of Reading) & Isobionics

Development of novel bacterial strains for production of hydrophobic compoundsPaul Herron (University of Strathclyde) & Ingenza

Regulation of methacrylate tolerance in E. coliIan Kerr (University of Nottingham) & Lucite International

Membrane selectivity and receptor recognition by chimeric antimicrobial peptidesBoyan Bonev (University of Nottingham) & DuPont

Touchy feely GM algaeMike Allen (Plymouth Marine Laboratory) & Algenuity

The projects have recently concluded so look out for the case studies on our website: http://cbmnetnibb.group.shef.ac.uk/outputs/case-studies/proof-of-concept-funding/

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Vacation Scholarships

20 undergraduate students19 academics

13 institutions3 companies

£34,000 from CBMNet£3,000 in-kind from industry

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Market analysis of the IBBE sector: relevant areas to Project DETOXDave Kelly, University of Sheffield & Project DETOX

The ChallengeProduct toxicity is a serious barrier to commercialization of Industrial Biotechnology and Bio-Energy (IBBE). Project DETOX is a BBSRC-funded collaboration between academics (at York, Nottingham, Sheffield, Cambridge and Exeter) and UK-based biotechnology companies. It aims to understand how microbes overcome product toxicity and to generate solutions for use in bioproduction.

Beyond the scope of the initial 5-year project, the DETOX team wanted to know who might be interested in their technology with a view to future commercialisation. In addition, they also wished to develop case studies for promotional purposes, based on feedback from industrial partners.

The ResearchProfessor Dave Kelly is a Professor of Microbiology at the University of Sheffield. The research in his laboratory focuses mainly on the physiology of bacterial pathogens and industrially relevant bacteria. Professor Kelly is also a collaborator in the DETOX project that aims to develop solutions to industrial biomanufacturing constraints.

Professor Kelly applied for a CBMNet Vacation Scholarship in order to fund important market research for the DETOX project and to develop promotional materials. Connor Innes, an undergraduate from the University of Sheffield, was employed as a CBMNet-funded summer student performing market research for the DETOX project.

The ResultConnor compiled a spreadsheet of companies working on biobased technologies, collecting pertinent data about the company and/or technology. Connor focused on bulk chemicals, where improving the yield of a biomanufacturing route has the potential to make the biggest difference to the economic viability of a biobased product.

Connor also conducted interviews with industrial partners of the DETOX project. Feedback was gathered on how the industrial partner valued their involvement with the DETOX Project and opportunities for working together beyond the project. This was used to prepare case studies and compiled into a leaflet that can be put on the website and used for marketing to new clients.

Connor presented his database and promotional leaflet to the DETOX team’s quarterly meeting.

The FutureThe spreadsheet of potential clients is a valuable resource for the DETOX project and will be updated with new companies as these emerge. It is an excellent starting point to develop a commercialisation strategy.

The meetings with industrial partners were useful to understand their perspective and how new clients may want to work with the DETOX project.

The case studies are essential to demonstrate to potential clients the benefits of working with Project DETOX. These promotional materials will be developed and published on the DETOX website and/or shared with clients and investors.

Connor gained experience of market analysis and presentation skills and will start a graduate job in September.

“Working with the DETOX project gave me in-depth exposure to bioindustry and greatly improved my understanding of the commercialisation of science, as well as offering the chance to improve soft skills such as presenting”

Connor Innes, Summer Student, University of Sheffield

“It has been excellent having Connor join the team to perform this essential market analysis. His work will directly contribute to an informed business plan to commercialise the technology we are developing on DETOX”

Joyce Bennett, DETOX Project Manager

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Protein-mediated transport across hybrid lipid-block copolymer membranesLars Jeuken, University of Leeds

The ChallengeMembrane proteins have a wide range of potential applications in industry. However, their application can be limited by the short functional lifetime of expressed proteins, particularly when reconstituted into liposomes.

Previous research has shown that reconstituting membrane proteins in hybrid lipid-block copolymer vesicles can increase their functional half-life by an order of magnitude. Building on this, the challenge is to develop new methodologies for the purpose of enhancing the stability and/or longevity of membrane proteins.

The ResearchLars Jeuken is a Professor of Molecular Biophysics at the University of Leeds. The research in his laboratory aims to developed novel biophysical and biotechnological tools for membrane proteins.

Professor Jeuken applied for a CBMNet Vacation Scholarship Award to fund an undergraduate student to carry out research in his laboratory. The project aimed to develop new methodologies to create hybrid polymer-lipid-protein vesicles from polymersomes and bacterial inner membrane extracts. The ultimate goal is to enhance the stability and/or longevity of membrane enzymes.

The ResultThis research project tested various methods to induce the formation of hybrid lipid-polymer vesicles, including tip-sonication, freeze-thaw and destabilisation of polymersomes with Triton X-100. Of these, only detergent-destabilisation with Triton X-100 was successful in forming hybrid vesicles from bacterial inner membrane extracts and polymersomes.

Hybrid vesicles were analysed by density gradient ultracentrifugation, showing that the ratio between polymer and inner membrane was heterogeneous. Importantly, the hybrid vesicles contained an active membrane-bound hydrogenase from the bacterial inner membrane.

The FutureIn future experiments, it will be important to assess whether hybrid lipid-polymer-inner membrane vesicles are able to enhance the stability of the membrane-bound hydrogenase from the inner membrane extract. It will also be essential to determine whether recombinant proteins reconstituted into hybrid vesicles can orientate and function correctly.

The data generated in this project will be used as pilot data or preliminary data to support a BBSRC responsive mode proposal. The undergraduate student gained valuable practical laboratory experience in an IB-focused field and, in the future, would like to pursue a PhD in a related area.

“It was fortunate to get this scholarship, taken up by one of our most talented undergraduate students. The project was challenging, but ultimately successful, providing valuable pilot data to guide our future research in biotechnology”

Prof Lars Jeuken, University of Leeds

“The scholarship I received helped me to develop my skills and gave me insight in a mysterious and wonderful world of science”

CBMNet Vacation Student

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Durable vesicles for stabilisation of membrane proteins in biotechnologyPaul Beales, University of Leeds

The ChallengeThe Beales laboratory are developing advanced vesicular materials as enhanced reconstitution systems that improve their stability and functional lifetime of membrane proteins. Many challenges remain in understanding the mechanism of enhanced functional stability, optimising materials and protocols and exploring a wider range of membrane proteins in these systems.

The ResearchThe Beales laboratory use cytochrome bo3 as their model membrane protein for development of hybrid vesicle technology; quantitative functional assays exist that allow easy comparison of the efficacy of different material compositions and reconstitution methods. Functional activity is measured by spectroscopically monitoring ubiquinol oxidation and, during this project, an electrochemical method to characterise oxygen reduction was introduced to monitor turnover of both substrates. The functional output of proton transport is measured by a pH-sensitive fluorophore, a strategy developed in this project.

Towards the development of hybrid vesicles as a robust and stable reconstitution system for membrane proteins, the Vacation Scholarship aimed to characterise proton pumping of cyt bo3 in these systems and investigate whether an alternative triblock copolymer (PMOXA-PDMS-PMOXA) could be substituted for our usual PBd-PEO diblock copolymer for successful functional reconstitution of the protein.

The ResultAn alternative triblock copolymer was successfully developed for functional reconstitution of cyt bo3 in hybrid vesicles, expanding our current parameter space for these systems and giving greater flexibility for future material optimisation.

The project found that PBd-PEO vesicles and their hybrids with lipid POPC had much lower passive proton permeability than liposomes. However reconstituted proteins only showed small ensemble pH shifts inside these vesicles suggesting they were fairly randomly oriented after reconstitution. The enzyme showed good functional activity when reconstituted using the new triblock copolymer, opening a further avenue for future investigation.

The student successfully reproduced previous work by functionally reconstituting cyt bo3 in hybrid vesicles and quantifying their functional activity by spectroscopically monitoring the oxidation of ubiquinol. Furthermore, she used a new functional activity by electrochemically monitoring the reduction of oxygen to confirm enzyme activity by a second method.

The student measured the proton permeability of lipid, polymer and hybrid vesicles to protons, finding polymer and hybrid membranes had very low proton permeability. Proton pumping assays reported by HPTS fluorescence showed small (<0.1 pH unit) shifts inside vesicles in ensemble experiments, strongly suggestive of the proteins being fairly randomly oriented within the membranes.

The student successfully reconstituted cyt bo3 in vesicles using the alternative PMOXA-PDMS-PMOXA triblock copolymer, expanding our material parameter space for further study. We did not have time to investigate SMALP reconstitution methods. As anticipated, this was ambitious within the time available and so F-ATPase was not investigated.

The FutureThe Vacation Scholarship helped consolidate the Beales Lab newly established hybrid vesicles method and demonstrate feasibility of some new materials and methods within this framework. The data will contribute to preliminary data for future grant applications and may contribute to a submission to Methods journal on the hybrid vesicle technique.

In the immediate term this work will be carried forward by a first year PhD student funded through EPSRC SOFI CDT; the primary focus of this student will be characterising the physical properties of hybrid membranes and correlating this to the biochemical properties of enhanced membrane protein durability in these environments to better understand the stability mechanism and guide further optimisation.

We are also seeking to recruit a biochemistry PhD student through the BBSRC White Rose DTP, who would explore the properties of a wider range of membrane proteins reconstituted in hybrid vesicles and develop optimised methods for their reconstitution.

“CBMNet Vacation Scholarships are a great scheme for getting talented undergraduates into the lab to gain valuable research experience while making important contributions to ongoing projects.”

Dr Paul Beales, University of Leeds

“Overall this experience has been really enjoyable and has helped me confirm that I would like to continue studying past my undergraduate degree.”

Ellen Moscrop, CBMNet Vacation Student

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Functional characterisation of a putative succinate efflux pump from Corynebacterium glutamicumChris Mulligan, University of Kent

“I was very pleased to be awarded the CBMNet Vacation Scholarship as it allowed my lab to start a completely new project on a very interesting, but poorly understood, family of transporters. The materials produced and the information obtained from this project will be very useful for future studies on these transporters.”

Dr Christopher Mulligan, University of Kent

“Before starting the project, I had a vague plan of pursuing laboratory based science as my career and since doing my summer project I am now confident that doing a PhD is my future career plan. I learnt more lab skills within three weeks of this project than I have in two years of teaching labs and I am forever grateful for that. The only disappointing aspect was that the I didn’t want the project to end and have to go back to attending lectures!”

Alice Evans, Vacation Student, University of Kent

The ChallengeSuccinate is a key precursor in the production of biodegradable plastics and fabrics. The majority of industrially produced succinate is derived from petrochemical precursors. However, several microbial species have been engineered to maximise succinate production during fermentation.

A succinate efflux pump, SucE, was recently identified in C. glutamicum, which substantially increases succinate production when overexpressed. However, the structure, mechanism, energetics and substrate specificity of this transporter remain unknown.

A comprehensive understanding of SucE’s transport mechanism could allow us to manipulate this transporter and/or its energy source to make efflux of succinate (and possibly other dicarboxylic acids) more efficient, potentially increasing the succinate yield of C. glutamicum.

The first step on this path is to develop an expression system with which to overexpress and purify SucE in sufficient quantities for biochemical characterisation.

The ResearchDr Chris Mulligan is a Lecturer in Molecular Biosciences at the University of Kent. His research interests centre on understanding the molecular mechanism of membrane transporters using a range of biochemical and biophysical methods.

Dr Mulligan applied for a CBMNet Vacation Scholarship that allowed him to train an undergraduate student in his lab in the overexpression and purification of integral membrane proteins for biochemical characterisation.

The undergraduate student, Alice Evans, was trained in general molecular biology and biochemical techniques, including gene cloning, microbial culturing, recombinant protein expression, affinity purification, SDS-PAGE and Western blotting. Alice also received training in techniques specific to membrane proteins, including membrane vesicle preparation and detergent solubilisation.

The ResultUnder the supervision of Dr Mulligan, Alice amplified SucE from C. glutamicum and cloned the gene into bacterial expression vectors. Alice systematically screened protein expression under a series of conditions, and purified the protein using immobilised metal affinity chromatography.

While the quantity of protein produced was not sufficient for further characterisation, this work lays the foundation for further expression optimisation in the future.

The FutureThis CBMNet Vacation Scholarship-funded research project provided excellent training and a research lab experience for Alice in the characterisation of membrane proteins. This project has strengthened Alice’s desire to pursue a career in research science starting with post-graduate training after the completion of her undergraduate degree.

This project has provided Mulligan lab with important information regarding the overexpression of SucE. The encouraging results produced from this project mean that the characterisation of succinate efflux pumps, including SucE, will remain a major research project in the Mulligan lab in the future.

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Examining the potential for periplasmic volume manipulation and its influence on Sec and FT3SS based protein productionGraham Stafford, University of Sheffield & FUJIFILM Diosynth Biotechnologies

“It is always a pleasure to host Undergrad students on summer projects, I know it kick-started my career and I gained great value from the experience.”

Graham Stafford, University of Sheffield

“I thoroughly enjoyed the project, and being part of novel research. I also experienced the joys as well as sometimes frustrating realities of real science.”

Jack Hales, Vacation student, University of Sheffield

The ChallengeE. coli is a vital organism to industrial biotechnology and is currently used for the production of a range of important biopharmaceutical proteins, such as insulin. Recent work has suggested that the volume of the periplasmic space may be important in protein production in E. coli and that increasing periplasmic volume may alleviate envelope stress responses resulting from protein production.

The aim of this summer vacation studentship was to modify the E. coli periplasmic volume by altering the length of Braun’s Lipoprotein (Lpp) and assess what effect this has on protein production in the periplasm (Sec-dependent secretion) and on one-step FT3SS-mediated secretion.

The ResearchDr Graham Stafford is a Reader in Microbiology at the University of Sheffield. The research in his laboratory focuses mainly on capitalising and using molecular microbiology knowledge to combat disease pathogens or for biotechnology applications.

FUJIFILM Diosynth Biotechnologies (FDB) is a world leading company involved in the development and production of biologics, vaccines and advanced therapies.

Dr Stafford applied for a CBMNet Vacation Scholarship to fund an undergraduate student to carry out research in his laboratory. The project aimed to engineer E. coli to improve IB processes for protein production in bacteria.

The ResultIn order to alter the periplasmic space of E. coli, the vacation student successfully generated a deletion strain that no longer expressed Lpp. The next step was to design and synthesise plasmids that would express mutant Lpp genes with altered lengths, in order to complement the deletion strain. These plasmids have been successfully generated.

As a model, a protein of interest to FDB was used to test expression in the systems under study.

Despite not completing every aim the student (Jack Hales) gained valuable experience of molecular biology, visiting the FDB facility and being embedded in a University research department.

The FutureThe next steps in this project will need to generate E. coli strains that express lengthened or shortened Lpp proteins and to test these strains for protein production. It will be interesting to see how periplasmic volume affects protein production via the two secretion pathways and if increasing/reducing periplasmic volume has different effects on these.

This vacation scholarship has laid the groundwork for further research on this project and it will be taken forward by a fourth year MBiolSci student in the next year, hopefully to form the basis of further funding applications. It is also closely aligned with current research collaborations between Dr Stafford’s laboratory and FDB, strengthening their academic-industrial partnership.

The undergraduate student gained some great, real-life experience in a research laboratory and also visited the FDB facilities.

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Spotlight on Industry

28 different companies£107,000 cash contribution

£455,000 in-kind contribution

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Dr Jen VanderhovenVice President Global Business Change, FUJIFILM Diosynth Biotechnologies (FDB)

What is your background and current job role?As Vice President Global Business Change, I am responsible for developing the FDB growth strategy to ensure we meet our global vision, mission and business objectives. I ensure successful implementation of strategic initiatives and lead the Executive Leadership delivery planning to transform the business and meet the challenges of efficient and sustainable growth. I also oversee the day to day running of our Global Project Management Office, ensuring we have a global change delivery capability that is sufficient to support the activities required for long term business success.

Educated to PhD level in the field of biochemistry, I have a 12-year progressive career spanning both the commercial and higher education sectors, with significant experience in realizing challenging strategic visions and operational goals within large and complex environments. I have previously held roles in Business Development, Research Development, Sales and Marketing, and Management positions in the Manufacturing sector. My previous role before FDB was actually as CBMNet Manager!

What Industrial Biotechnology and Bioenergy related project is currently being undertaken by your organisation?Fujifilm Diosynth Biotechnologies (FDB) provides process development and manufacturing services to the biopharmaceutical industry. Innovation activities take place on all three FDB sites (UK, North Carolina and Texas). In the UK our site at Billingham, is home to more than 250 scientists and engineers supporting activities including microbial and

mammalian cell line construction, biologics process and analytical development and extensive characterisation services for products and processes that we operate for our customers. We believe continued innovation and adoption of new approaches and technologies have been key to our success, and will continue to be essential as we strive to realise our goal ‘to be the leading and most trusted global CDMO partner in the biopharmaceutical industry’.

What do you think the challenges related to this project are in the next 1-5 years?Our cross-site global innovation activities cover many disciplines and include development of potentially disruptive technologies as well as incremental improvements to existing technology, leading to a defined step change.

FDB has three overarching FDB innovation themes that we see as essential for development in the coming years, based on a review of market and business needs: Dry Science, to use automation, robotics and predictive technologies to reduce the amount of wet, bench science required to meet project objectives; Manufacturing 2.0 to develop next generation manufacturing & testing technologies using smaller, more flexible, more automated facilities that are aligned across sites; New Biology, to support emerging product types, synthetic manufacture and step out science.

How will the contacts made via CBMNet help you/your organisation in the future?The UK’s strong science base in biopharmaceuticals and industrial biotechnology provides many groups who can support and augment our in-house innovation efforts. Much of our innovation work is driven and resourced internally, or in collaboration with our suppliers or customers. However, FDB have historically also supported a significant number of external innovation projects including supporting PhD studentships, participating in Innovate UK-funded consortium projects, and stand-alone fee for service work with third party companies. Many of these contacts we have made through CBMNet activities.

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What is your background and current job role?I have a BSc (Hons) in Applied Biology from Newcastle University. I have worked at Fujifilm Diosynth Biotechnologies and its predecessors MSD and Avecia for 15 years. I began in a lab-based role delivering strain construction aspects of customer projects and working on internal technology development projects, one of which led to the successful pAVEway expression system. Over time my role has changed to become less lab-based and more office-based. I currently manage the Molecular & Micro Biology team of four lab based Scientists and supervise the delivery of early strain construction Process Development projects on behalf of our clients, in addition to supervision of external collaborations at partner Universities and involvement in the CBMNet.

What Industrial Biotechnology and Bioenergy related project is currently being undertaken by your organisation?We have several long-term collaborations which have stemmed from contacts made at CBMNet events and initiated with CBMNet Proof of Concept and Business Interaction Vouchers. We have just started a ‘Stand-Alone’ LINK award with Dan Mulvihill at Kent University; are about six months into a CASE Studentship with Graham Stafford at Sheffield University; and are coming to the end of two PhD awards at Edinburgh University with Teuta Pilizota and Louise Horsfall.

What do you think the challenges related to this project are in the next 1-5 years?The biggest challenges with all of these projects will be to turn something which has shown promise at small scale, in shake flask or 250 mL bioreactor with one or two proteins into a technology which can be used robustly on an industrial process with tangible benefits at 5,000 L scale or larger.

How will the contacts made via CBMNet help you/your organisation in the future?The CBMNet has introduced me to many contacts with interesting technology, who’s paths I otherwise would not have crossed. The funding available has enabled collaborative projects to start which will hopefully lead to the development of technologies that can be put to use to enable the economical manufacture of pharmaceuticals to the ultimate benefit of patients.

Dr Chris LennonStaff Scientist (Molecular Biology), FUJIFILM Diosynth Biotechnologies

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And many thanks to our ever-supportive Management Board...

Dr Alan GoddardAston University

Dr Chris LennonFujifilm Diosynth Biotechnologies

Dr Sam MillerUniversity of Aberdeen

Prof Susan Molyneux-HodgsonUniversity of Exeter

Dr Anna Sobolewska-StawiarzCroda

Prof Gill StephensUniversity of Nottingham

Dr John HeapImperial College London

Prof Lars JeukenUniversity of Leeds

Prof Dave KellyUniversity of Sheffield

Dr Preben KrabbenMicrobesphere Ltd.

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Looking Ahead

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Algae-UK: Exploiting the algal treasure troveSaul Purton, University College LondonCo-director(s): Patricia Harvey, University of Greenwich; Michele Stanley, Scottish Association for Marine Science; Anna

Amtmann, University of Glasgow

Biomass Biorefinery Network (BBNet)Simon McQueen-Mason, University of YorkCo-director(s): Michele Stanley, Scottish Association for Marine Science; Elizabeth Thornley, Aston University; Dimitrios

Charalampopoulos, University of Reading; David Leak, University of Bath

High Value Biorenewables (HVB) NetworkIan Graham, University of YorkCo-director(s): Joe Ross, University of York, Anne Osbourn, John Innes Centre

Carbon Recycling: Converting waste derived GHG into chemicals, fuels and animal feed (CCNet)

Professor Nigel Minton, University of NottinghamCo-director(s): Sonia Heaven, University of Southampton; Saul Purton, University College London; Mark Poolman, Oxford

Brookes University; Brigitte Nerlich, University of Nottingham; William Zimmerman, University of Sheffield

Environmental Biotechnology Network (EBNet)Sonia Heaven, University of SouthamptonCo-director(s): Thomas Curtis, Newcastle University; Frederic Coulon, Cranfield University; Tony

Gutierrez, Heriot-Watt University; Jhuma Sadhukhan, University of Surrey

Elements of Bioremediation, Biomanufacturing & Bioenergy (E3B) Metals in Biology

Nigel Robinson, Durham UniversityCo-director(s): Martin Warren, University of Kent; Jonathan Lloyd, University of Manchester

Networks in Industrial Biotechnology & BioenergyPhase II: looking forward to the future

@EBNetUK

@HVB_net

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Microbial Cell Factory

Manipulation of substrate uptake

Optimisation of elux

Socio-economic challenges

Hijacking transporters for IBBE

Modifying the membrane

Putting it all together: consolidated bioprocessing

Testing novel mechanisms of protein secretion for use in biotechnology

Identifying e�lux systems for commodity chemicals

Identification and characterisation of e�lux pumps for the important

feedstock, succinate

Understanding the e�ects of solvents on bacterial cell membranes in order to optimise n-butanol production

Designing mimetic bacterial membranes as tools for the development of antimicrobial agents

Improving the tolerance of microbes to toxic solvents

Development of an improved screen for intracellular toxicity

Optimising membrane function in the Clostridial ABE process

Tools to generate new Clostridium strains for IB

Optimising protein production by secretion into recombinant outer membrane vesicles

Screen to identify bottlenecks in recombinant protein secretion during industrial processes

Development of new methodologies to isolate optimal strains for IB

Novel rechnologies for characterising membrane proteins

Engineering yeast to use waste as a feedstock to produce valuable products

Identification of membrane transporters that will allow bacteria to utilise lignin monomers

Characterisation of transporters involved in human malodour production

Market Analysis of the IBBE Sector

Training events and career progression for Eearly Career Researchers

Public art work - Festival of the Mind

CBMNet Event: Social and Political Challenges for the Bioeconomy

Moving complex molecules across membranes

Engineering bacteria and yeast strains to express biosurfactants

Engineering of E. coli to optimise antibody production

Polysaccharide production by microalgae

Studying the e�ects of osmolarity on E. coli and how this impacts on protein production

CBMNet Themes - Our Legacy

39

Microbial Cell Factory

Manipulation of substrate uptake

Optimisation of elux

Socio-economic challenges

Hijacking transporters for IBBE

Modifying the membrane

Putting it all together: consolidated bioprocessing

Testing novel mechanisms of protein secretion for use in biotechnology

Identifying e�lux systems for commodity chemicals

Identification and characterisation of e�lux pumps for the important

feedstock, succinate

Understanding the e�ects of solvents on bacterial cell membranes in order to optimise n-butanol production

Designing mimetic bacterial membranes as tools for the development of antimicrobial agents

Improving the tolerance of microbes to toxic solvents

Development of an improved screen for intracellular toxicity

Optimising membrane function in the Clostridial ABE process

Tools to generate new Clostridium strains for IB

Optimising protein production by secretion into recombinant outer membrane vesicles

Screen to identify bottlenecks in recombinant protein secretion during industrial processes

Development of new methodologies to isolate optimal strains for IB

Novel rechnologies for characterising membrane proteins

Engineering yeast to use waste as a feedstock to produce valuable products

Identification of membrane transporters that will allow bacteria to utilise lignin monomers

Characterisation of transporters involved in human malodour production

Market Analysis of the IBBE Sector

Training events and career progression for Eearly Career Researchers

Public art work - Festival of the Mind

CBMNet Event: Social and Political Challenges for the Bioeconomy

Moving complex molecules across membranes

Engineering bacteria and yeast strains to express biosurfactants

Engineering of E. coli to optimise antibody production

Polysaccharide production by microalgae

Studying the e�ects of osmolarity on E. coli and how this impacts on protein production

Get in touch...

CBMNetUniversity of SheffieldFirth CourtWestern BankSheffield S10 2TN

http://www.cbmnetnibb.net/

0114 222 2745

cbm@sheffield.ac.uk

@CBMNet_NIBB

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