Protein Expression Facility (PEF)

The Protein Expression Facility (PEF) provides a recombinant expression and purification service to the University of Birmingham Research community.

As part of the Technology hub the PEF acts as a focal point for advice about all aspects of recombinant protein production. The expression systems used by the PEF will enable the recombinant proteins generated to be used in a wide range of downstream applications.

The PEF provides a service as close to cost price as possible, but consistent with facility sustainability. The PEF aims to produce a wide variety of proteins to enable a comprehensive range of downstream technologies and to be responsive to customer needs where possible.

The PEF resides within a designated laboratory suite that was funded by a central infrastructure bid in 2010 through the College of Medical and Dental Sciences.

The PEF is self-funded, with annual running costs met through recharging schemes proportional to the level and nature of collaboration. Activities are distributed across the UoB research community, but the PEF welcomes collaboration with Midlands Innovation and external companies.

The PEF provides a central location to undertake recombinant protein expression within a dedicated environment. It also provides a range of expression and purification services or training which can be commissioned at any time.

Protein Expression Facility Team

Protein Expression Specialist

Jamie R. M. Webster

Jamie joined the Protein Expression facility in 2013. He was awarded a BSc (Hons) in Applied Science (1986) from the Polytechnic, Wolverhampton, a MSc in Science (F12 - frontiers in medical science) with Distinction from the Open University (2009) and holds a current RSci from the Science Council (IST 2015-2020, RSB 2020 to date). Jamie has thirty-six years technical experience both with the civil service (MAFF), NHS (NBTS) and HEI’s (Cambridge, Nottingham and Birmingham). His experience prior to joining the facility supported research in Cyanobacterial proteins and Photosystem II, Archaeal Cas3 and Cas5, the Chromosomal passenger complex and Wallerian Degeneration and since joining the facility has supported colleagues in the college of MDS, LES and School of Chemistry.

Email: J.R.M.Webster@bham.ac.uk

Research Facilities Manager

Dr Adriana Flores-Langarica

Email: A.FloresLangarica@bham.ac.uk

Protein Expression Academic Leads

Professor Benjamin Willcox

Email: B.Willcox@bham.ac.uk

Professor Steve Smerdon

Email: S.J.Smerdon@bham.ac.uk

Training and Support

Equipment in the Protein Expression Facility (PEF)

We arrange and co-ordinate training for users to independently use the AKTA FPLC, PHERAstar and the Biacore T200. Currently our training programmes are divided in two sections.

Firstly, for the PHERAstar FS contact the specialist to add you to the Technology Hub Canvas module which can be completed at the user pace and once that is completed, we can deliver an in-person training session at the equipment. If you require training just request it on the Stratocore login page for UoBPEF. We deliver sessions to suit customer need.

Secondly, for AKTA FPLC the we can deliver an in-person training session at the equipment. If you require training just request it on the Stratocore login page for UoBPEF. We deliver sessions to suit customer need.

Finally for the Biacore T200 SPR we arrange an in-person training session for groups of up to six people with the manufacturer specialist team at the equipment which is charge to the user as this is a specialist piece of equipment. If you require training, please contact pef@contacts.bham.ac.uk for current prices for the specialist training and to arrange timings.

We deliver a full service for recombinant protein expression and purification, and this will be tailored to needs of the project undertaken. A quotation is provided setting out the phases and recharging process and once agreed between parties the work commences. Protein expression and purification training can be requested on a 1-2-1 basis if mentoring is required for a specialist piece of work. For information and price on application please contact pef@contacts.bham.ac.uk.

Access and Booking

External users and price on application internal users are encouraged to contact us via email in the first instance. For the AKTA FPLC, T200 and PHERAstar all internal prospective users need to request an account on the core facilities booking system Stratocore (unless you already have one). Please ensure you use your UoB email address to register your Stratocore account.

As part of your account, you will need a valid finance/grant code. We need this to be able to charge you for your use of the equipment.

Your code will either be a project or general ledger. Guidance on finance/grant codes is here:

Project Ledger. (Project Finance Number begins with a ‘1’ or ‘2’). Project Ledger codes require 3 components in the format:

Project Ledger Code Components
Cost Centre Code	Task Number	Project Finance Number
C082	1.1	21991

General Ledger. (Project Finance Number begins with a ‘6’). General Ledgers require 2 components in the format:

General Ledger Code Components
Cost Centre Code	Project Finance Number
C283	65371

If you are unsure about any of your finance codes, contact techubfinance@contacts.bham.ac.uk who will be able to assist you.

Equipment and Services

The PEF offers a full service in the three expression models and downstream purification of proteins but will also offer selected service provision to cater for customer needs. Examples include:

Expression construct design and bioinformatics
Dedicated culture facilities for prokaryotic and eukaryotic expression
Expression systems available: E.coli, mammalian cell and insect cell expression
Fast Protein Liquid Chromatography
PHERAstar FS microtiter plate reader
Affinity purification
MHC Class I Monomer production
Training in purification techniques
Endotoxin testing

Expression and Purification Facility

The PEF offers a one stop shop for small scale test expressions through to medium scale expressions followed by purification and polishing of the protein requested.

The service provided may be inclusive of cloning, optimisation of the construct via test expression through to scaling up for purification and polishing, or may be a Principal Investigators well characterised protocol. Work undertaken will depend on the initial discussions to outline the project requirements and the status of existing Biosafety approvals.

The PEF offers expertise in E.coli, mammalian cell and insect cell expression and has two dedicated culture facilities. One is dedicated to prokaryotic work and has three programmable incubators capable of growth conditions between 4-80 degrees with two dedicated Beckman Avanti centrifuges. The eukaryotic culture facility hosts incubators for static, shaking and roller bottle culture of mammalian or insect cell.

Downstream applications of refolded MHC1 Protein table, Top to bottom: Crystallographic Structure Determination, Surface Plasmon Resonance Studies, Circular dichroism (CD) measurements and flow cytometry

MHC class I refolds

Biotinylated MHC Class I Monomer are composed of a particular peptide in combination with a particular MHC class I allele that has been biotinylated at the C-terminus of the heavy chain extracellular domain. The Biotinylated Class I MHC/peptide complex can be used in research studies using the MHC tetramer assay and the PEF has been routinely supplying the Birmingham Researcher with monomers since 2010.

The PEF routinely makes HLA A0101, HLA A0201, HLA B8 and HLA B7 and holds a supply of the inclusion bodies used to refold the peptides chosen by the customer.

For more information regarding how to get in touch with the PEF about Biotinylated MHC Class I Monomer production, please use the contact details at the bottom of this webpage.

Surface Plasmon Resonance - Biacore T200

Access to a Biacore T200 is available for internal and external customers to use with booking via ppms. The equipment is run as a user operated service and training can be arranged for researchers.

For more information regarding how to get in touch with the PEF about the Biacore T200 and for any training requests, please use the contact details at the bottom of this webpage.

Acknowledgement Statement

The University of Birmingham (UoB) is a founding signatory of ‘The Technician Commitment’. A sector wide, collaborative initiative that aims to ensure visibility, recognition, career development and sustainability for technical staff working in higher education and research, across all disciplines.

The UoB has invested and supports the Technology Hub facilities not only with equipment but also specialist staff. The staff are a key element of our offering since they not only ensure that all equipment is looked after and performing to specification, but they also have a significant experience, and the input and advice is very valuable to our users.

In line with this, the Technology Hub Facilities request that all users of the facilities to recognize its contribution to manuscripts in one of the following ways:

The authors would like to acknowledge the (Facility) at the UoB for support of (technology used) experiments. The authors would like to acknowledge (specialist name). In some instances, the specialist may make substantial contributions to the conception / design of the work or the acquisition, analysis, or interpretation of data for a manuscript. In this case, it is expected that this is appropriately recognized through co-authorship on any publications resulting from the work. Criteria for inclusion of a specialist as an author on a publication. Where the specialist has either:

Performed experiments or analysis, the data from which is included in the manuscript.
Produced customized analysis workflows or scripts for the work.
Significantly contributed to the design, conception, or interpretation of the experiments.

Recent Publications

Eldershaw, S.A., Pearce, H., Inman, C.F., Piper, K.P., Abbotts, B., Stephens, C., Nicol, S., Croft, W., Powell, R., Begum, J., Taylor, G., Nunnick, J., Walsh, D., Sirovica, M., Saddique, S., Nagra, S., Ferguson, P., Moss, P. and Malladi, R. (2021), DNA and modified vaccinia Ankara prime–boost vaccination generates strong CD8+ T cell responses against minor histocompatibility antigen HA-1. Br J Haematol. https://doi.org/10.1111/bjh.17495

Odintsova, E., Mohammed, F., Trieber, C. et al. Binding of the periplakin linker requires vimentin acidic residues D176 and E187. Commun Biol 3, 83 (2020). https://doi.org/10.1038/s42003-020-0810-y

Pedrera M, Macchi F, McLean RK, Franceschi V, Thakur N, Russo L, Medfai L, Todd S, Tchilian EZ, Audonnet JC, Chappell K, Isaacs A, Watterson D, Young PR, Marsh GA, Bailey D, Graham SP, Donofrio G. Bovine Herpesvirus-4-Vectored Delivery of Nipah Virus Glycoproteins Enhances T Cell Immunogenicity in Pigs. Vaccines (Basel). 2020 Mar 2;8(1). pii: E115. doi: 10.3390/vaccines8010115.

Mohindar M.Karunakaran, Carrie R.Willcox, Mahboob Salim, Daniel Paletta, Alina S.Fichtner, Angela Noll, Lisa Starick, Anna Nöhren, Charlotte R.Begley, Katie A.Berwick, Raphaël A.G.Chaleil, Vincent Pitard, Julie Déchanet-Merville, Paul A.Bates, Brigitte Kimmel, Timothy.J.Knowles Volker Kunzmann, Lutz Walter and Thomas Herrmann. Butyrophilin-2A1 Directly Binds Germline-Encoded Regions of the Vγ9Vδ2 TCR and Is Essential for Phosphoantigen Sensing Immunity Volume 52, Issue 3, 17 March 2020, Pages 487-498.e6 https://doi.org/10.1016/j.immuni.2020.02.014

Fiyaz Mohammed, Daniel H. Stones Benjamin E. Willcox, Application of the immunoregulatory receptor LILRB1 as a crystallisation chaperone for human class I MHC complexes, Journal of Immunological Methods, Volume 464, January 2019, Pages 47-56, https://doi.org/10.1016/j.jim.2018.10.011

Jamshad M, Knowles TJ, White SA, Ward DG, Mohammed F, Rahman KF, Wynne M, Hughes G1, Kramer G, Bukau B, Huber D. The C-terminal tail of the bacterial translocation ATPase SecA modulates its activity, Elife. 2019 Jun 27;8. pii: e48385. doi: 10.7554/eLife.48385

Willcox CR, Vantourout P, Salim M, Zlatareva I, Melandri D, Zanardo L, George R, Kjaer 4, Jeeves M, Mohammed F, Hayday AC, Willcox BE. Butyrophilin-like 3 Directly Binds a Human Vγ4+ T Cell Receptor Using a Modality Distinct from Clonally-Restricted Antigen. Immunity. 2019 Nov 19;51(5):813-825.e4. doi: 10.1016/j.immuni.2019.09.006. Epub 2019 Oct 15.

Davey, Martin S.; Willcox, Carrie R.; Hunter, Stuart; et al (2018) The human V delta 2(+) T-cell compartment comprises distinct innate-like V gamma 9(+) and adaptive V gamma 9(-) subsets .NATURE COMMUNICATIONS Open access indicator Volume: 9 Article Number: 1760 Published: MAY 2 2018

Hunter, S., Willcox, C., Davey, M., Kasatskaya, S., Jeffery, H., Chudakov, D., Oo, Y., Willcox, B., Human liver infiltrating γ δ T cells are composed of clonally expanded circulating and tissue-resident populations, Journal of Hepatology (2018), doi: https://doi.org/10.1016/j.jhep.2018.05.007