Professor Sheena Radford OBE, FMedSci, FRS

Introduction

One of the most fascinating questions in biology is how proteins are able to fold and assemble into complex, functional entities given just the information provided by the amino acid sequence. A related, equally important facet of the same fundamental question is how protein misfolding can lead to protein aggregation, cellular dysfunction and disease. These issues are the major focus of my research and are tackled using a broad range of techniques including protein chemistry, structural molecular biology, chemical biology, cell biology and biophysical methods.

Watch a lecture by Prof Radford discussing her work and celebrating her FRS award. Go to https://www.youtube.com/watch?v=r1eK3DLCMcM

Current major projects

• Mechanism(s) of protein misfolding and assembly into amyloid
• Outer Membrane Protein (OMP) folding – The role of chaperones & BAM
• Stabilising proteins of therapeutic interest against aggregation
• Method development (MS, NMR, single molecule, biophysical methods)

Detailed research programme

Mechanism(s) of protein misfolding and assembly into amyloid

How and why proteins aggregate into amyloid are important fundamental questions that have far-reaching biomedical importance. Focusing on the proteins β₂-microglobulin (dialysis-related amyloidosis); islet associated polypeptide (IAPP) (type II diabetes); α-synuclein (Parkinson’s) and Aβ (Alzheimer’s), work in our group aims to map the structural mechanism of amyloid formation and to develop reagents to control aggregation in vitro and in vivo. Recent highlights include structure determination of oligomers using NMR (Karamanos et al. (2019) eLife) and fibrils using cryoEM (with Neil Ranson) (Gallardo et al. (2020) Nature Struct. Mol. Biol.; Wilkinson et al. (2023) Nature Comms.), discovery of small molecules that modulate amyloid formation (Cawood et al. (2020) JACS, Xu et al. (2022) Nature Comms.); and demonstration that early protein-protein interactions in amyloid formation are specific and can be targeted to arrest assembly in vitro and in vivo (Doherty et al. (2020) Nature Struct. Mol. Biol., Ulamec et al. Nature Comms. (2022), Guthertz et al. PNAS (2022)).Working with Neil Ranson, we have also recently shown that amyloid fibril structures change with time (Wilkinson et al. (2023) Cell) and, with Rene Frank, have provided the first insights into amyloid in the mouse and human brain of Alzheimer’s patients (Leistner et al. (2023) Nat Comms; Gilbert et al. (2024) Nature).

Outer Membrane Protein (OMP) folding - The role of chaperones & BAM

How OMPs fold and assemble into the asymmetric outer membrane (OM) of Gram-negative bacteria is a second research theme in our lab. In a collaborative multidisciplinary team (with David Brockwell, Neil Ranson and Ian Collinson (Bristol)) we are investigating how OMPs cross the inner-membrane via SecYEG (Fessl et al. (2018) eLife; Crossley et al., (2024) EMBO J.); traverse the periplasm aided by chaperones (Skp/SurA) (Calabrese et al. (2020) Nature Comms), fold into membranes in vitro and in vivo catalysed by the beta-barrel membrane machinery (BAM) (Iadanza et al. (2020) Comms. Biol., Schiffrin et al. (2022) Comms. Biol.). Building on these insights we are currently exploring the dynamic motions of BAM during catalysis (Haysom et al., Angewandte Chemie) and how this can be harnessed to generate new antibacterial agents against Gram-negative pathogens (White et al. (2021) Nature Comms.). We have also worked with David Baker (Seattle) and Anastasia Vorobieva (Brussels) to design transmembrane beta barrel proteins and explore their folding and function in vitro (Vorobieva et al. (2021) Science; Berhanu et al., (2024) Science). Most recently, we have used smFRET and NMR methods to map the binding of client OMPs to their chaperone, SurA (Schiffrin et al. (2024) Nat. Comms) and used cryoEM, smFRET and proteomics to reveal the catalytic cycle of SurA caught in the act of delivering OMPs to BAM (Fenn et al. (2024) Nat Comms.).

Stabilising proteins of therapeutic interest against aggregation

We are also exploiting our knowledge of protein folding/aggregation to practical benefit by screening amyloidogenic proteins, as well as proteins of interest to biopharma, for hotspots that cause aggregation. By coupling aggregation to bacterial growth using a tripartite β-lactamase fusion construct we have discovered small molecules that prevent aggregation of amyloidogenic proteins (Saunders et al. (2016) Nature Chem. Biol.). With David Brockwell and our collaborators in AstraZeneca, we recently combined the assay with directed evolution to enhance the resilience of biopharmaceutically-relevant antibodies to aggregation (Ebo et al. (2020) Nature Comms.). Finally, in collaboration with David Brockwell and Nik Kapur (Mechanical Engineering, Leeds), we are examining how flow fields enhance, or cause, aggregation by flow-induced protein deformations (Dobson et al. (2017) PNAS, Willis et al. (2020) Eng. Rep.; Willis et al. (2024) J. Pharm. Sci.).

Method development (MS, NMR, single molecule, biophysical methods)

Major developments in methods and instrumentation have played a key role in increasing in our understanding of protein folding and aggregation. Future developments in these fields will also require innovative approaches that cross the boundaries between disciplines. We have been involved in many exciting collaborations to fulfil this aim. With Professors Alison Ashcroft (now emeritus) and Frank Sobott we have developed and expanded our arsenal of methods to interrogate protein folding, protein-protein interactions, and protein complexes, including MS methods (HDX-MS and fast photochemical oxidation of proteins (FPOP-MS)) (Cornwell et al. (2019) Analyt. Chem., Cornwell et al. (2021) J. Am. Soc. MS.); ion mobility MS to determine the effect of ligand binding on amyloidogenic monomers and oligomers (Byrd et al. (2023) J. Am. Soc. MS.) and, with Andrew Wilson (Chemistry), we have developed rapid crosslinking to map protein-chaperone interactions (Horne et al. (2018) Angewandte Chemie; Calabrese et al. (2020) Nature Comms.). Developments in NMR methods remain a mainstay of our laboratory activities (Karamanos et al. (2022) Frontiers Neurosci.). In collaboration with David Brockwell and George Heath we are involved in exciting developments in the use of the AFM for measurements of protein unfolding and protein binding (Ulamec et al. (2021) Nature Comms.) and with Paolo Actis (Mechanical Engineering) in the development of nanopores for the manipulation and identification of protein assemblies (Chau et al. (2020) Nanoletters; Chau et al. (2022) ACS Nano, Chau et al., (2024) Nat Comms.).

More information about all these projects can be found on the websites of our collaborators’ Astbury web pages.

For further details about the Radford laboratory, people involved, molecular images and available opportunities, please see http://sheena-radford-lab.uk