The Oxford Protein Production Facility-UK
Lead Research Organisation:
University of Oxford
Department Name: Structural Biology
Abstract
Proteins are essential components of all living cells. They comprise chains of amino acids coded for by specific DNA sequences and fold into three dimensional structures, that determine their function, for example as biological catalysts. Therefore, understanding protein structure and function is key to exploiting the information generated from sequencing of the human genome and in particular those of many human pathogens. Knowledge of individual proteins has made a major contribution to the development of a number of important drugs, e.g. Zanamivir (influenza neuraminidase inhibitor), Lapatinib (kinase inhibitor), Darunavir (HIV protease inhibitor). However, high throughput DNA sequencing continues to add to the database of known proteins at a rate that far exceeds our capacity to decipher their function. Therefore, accelerating the production of recombinant proteins is key to closing the gap between the acquisition of protein sequences and the determination of their structure and function. In response to this challenge, the Oxford Protein Production Facility will provide infrastructure to underpin the study of proteins on different length and time scales from atomic resolution (x-ray crystallography) to whole cell systems (optical microscopy).
Technical Summary
The Oxford Protein Production Facility-UK (OPPF-UK) is a National resource for protein science. The overall goal of the OPPF-UK is to provide the UK biological, chemical and biomedical communities with a world-class facility for the production of proteins for structural and functional research. The OPPF-UK has established a high throughput approach to the production of recombinant proteins using three main expression hosts (E. coli, mammalian cells and baculovirus/insect cells) which is provided as a service to the UK scientific community through a peer-review process. Research and development in the OPPF-UK will be focussed on streamlining the cloning, expression and purification of multi-protein complexes and membrane proteins using insect and mammalian cells. Methods will be rolled out as part of the user programme of the OPPF-UK. In parallel, data management systems will be enhanced to provide users with secure on-line access to all project data.
Planned Impact
Recombinant proteins play a key part in the drug discovery process, for example as components of biochemical screens for new chemical entities (NCEs), antigens for raising therapeutic antibodies and for crystallization to obtain structural information. We are contributing to projects involving the structure-based identification of NCEs, for example inhibitors of a novel metallo-beta-lactamase (NDM-1) which has been implicated in widespread antibiotic resistance of important bacterial pathogens. New knowledge will be added from these studies which may ultimately help in the development of new drugs. The OPPF-UK is developing a number of direct links with the biotech/pharma sector through consultancy and training and as a provider of protein production services. The OPPF-UK is also actively involved in helping to educate the next generation of potential research scientists. We contribute to school visits to the Rutherford Appleton Laboratory through staff giving talks about how proteins are studied in the OPPF-UK and our links with the Diamond Light Source. Finally, the OPPF-UK provides an important opportunity for staff development, since working in the OPPF-UK involves scientists in the management of multiple projects, dealing with deadlines and liaising with external users. This experience helps to develop transferable skills that are highly applicable not only to jobs in research & development but also other professions outside of science.
Publications
Ditsiou A
(2020)
The structure-function relationship of oncogenic LMTK3.
in Science advances
Byrne RT
(2013)
S-Adenosyl-S-carboxymethyl-L-homocysteine: a novel cofactor found in the putative tRNA-modifying enzyme CmoA.
in Acta crystallographica. Section D, Biological crystallography
Butryn A
(2020)
Molecular basis for GTP recognition by light-activated guanylate cyclase RhGC.
in The FEBS journal
Bird LE
(2014)
Application of In-Fusion™ cloning for the parallel construction of E. coli expression vectors.
in Methods in molecular biology (Clifton, N.J.)
Bird LE
(2015)
Green fluorescent protein-based expression screening of membrane proteins in Escherichia coli.
in Journal of visualized experiments : JoVE
Bird LE
(2016)
Expression Screening of Integral Membrane Proteins by Fusion to Fluorescent Reporters.
in Advances in experimental medicine and biology
Beale JH
(2015)
Crystal Structures of the Extracellular Domain from PepT1 and PepT2 Provide Novel Insights into Mammalian Peptide Transport.
in Structure (London, England : 1993)
Bayley CP
(2016)
Diversity between mammalian tolloid proteinases: Oligomerisation and non-catalytic domains influence activity and specificity.
in Scientific reports
Awanye AM
(2019)
Immunogenicity profiling of protein antigens from capsular group B Neisseria meningitidis.
in Scientific reports
Asiani KR
(2016)
SilE is an intrinsically disordered periplasmic "molecular sponge" involved in bacterial silver resistance.
in Molecular microbiology
Description | New001 Building research capacity for schistosomiasis drug discovery & development through high-content imaging & structural molecular biology studies |
Amount | £67,581 (GBP) |
Funding ID | MR/M026221/1 |
Organisation | Medical Research Council (MRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2015 |
End | 05/2017 |
Description | Structural Biology in the RFI |
Amount | £1,649,512 (GBP) |
Funding ID | EP/S025243/1 |
Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
Sector | Public |
Country | United Kingdom |
Start | 11/2018 |
End | 05/2020 |
Description | University of Oxford GCRF, HFCE |
Amount | £52,580 (GBP) |
Funding ID | 0006135 & 0005295 |
Organisation | Oswaldo Cruz Foundation (Fiocruz) |
Sector | Public |
Country | Brazil |
Start | 03/2018 |
End | 03/2019 |
Description | Building research capacity for schistosomiasis drug discovery and development through high-content imaging and structural molecular biology studies |
Organisation | Oswaldo Cruz Foundation (Fiocruz) |
Country | Brazil |
Sector | Public |
PI Contribution | Training in high throughput cloning and expression of proteins fro drug discovery research. Protein production and crystallization of Schistosoma proteins selected a potential drug targets. Delivery of seminar-based workshop at Oswaldo Cruz |
Collaborator Contribution | Expertise in drug screening Identification of potential inhibitors for co-crystallization experiments |
Impact | Technology transfer to Oswaldo Cruz. Novel results which will be published in due course. |
Start Year | 2015 |
Description | School visits |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Schools |
Results and Impact | Practical workshop for year 12 school groups ( approx. 50/visit ) on how recombinant proteins are produced using Green fluorescent protein as the example. |
Year(s) Of Engagement Activity | 2014,2015,2016 |
Description | Schools visit |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Schools |
Results and Impact | Hands-on practical workshops on proteins and protein crystallization |
Year(s) Of Engagement Activity | 2017 |
Description | Science Up Close: Rutherford Appleton Laboratory Open Day |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Public/other audiences |
Results and Impact | Hands-on crystallizing a protein for children as part of a stall about proteins and how we study them visited by attendees of the Rutherford Appleton Laboratory Open Day |
Year(s) Of Engagement Activity | 2015 |
Description | Science and Society Lecture |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Public/other audiences |
Results and Impact | Presented at the annual Science and Society lecture - talk sparked questions and discussions afterwords. Talk could lead to potential collaboration opportunists. |
Year(s) Of Engagement Activity | 2014 |
Description | Youtube video |
Form Of Engagement Activity | A broadcast e.g. TV/radio/film/podcast (other than news/press) |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | Video for general audience on protein production and crystallization produced by Backstage Science for the STFC. |
Year(s) Of Engagement Activity | 2012 |
URL | https://www.youtube.com/watch?v=joWNZDBqrlQ |