The Protein Choreography Group
In the Protein Choreography Group we identify mechanisms of dynamic functional control in protein molecules. We aim to understand the principles of allosteric regulation in biology and dysfunction in disease and to engineer protein dynamics for novel medicines and biotechnology tools. We develop interdisciplinary experimental and computational approaches, novel prototype instrumentation, software and mathematical models to achieve this. Our group has an international reputation for advancing millisecond time-resolved hydrogen/deuterium-exchange mass spectrometry (HDX-MS) methods to study large, dynamic molecular systems. Our new non-equilibrium methods allow us to see, for the first time, precisely how proteins reconfigure to activate, inactivate and to perform enzyme catalysis. Applied to natural systems, this yields insight into fundamental biology, including Parkinson’s disease pathogenesis, diabetes and cancers. Applied to medicine, this drives the design of switchable antibodies that activate inside the body and new screening technologies to discover safer and more effective classes of drugs.
In Parkinson’s disease and other synucleinopathies, the intrinsically disordered, presynaptic protein alpha-synuclein misfolds and aggregates. We hypothesise that the exposure of alpha-synuclein to different cellular environments, with different chemical compositions, pH and binding partners, together with post-translational modifications alter its biological and pathological function by inducing changes in molecular conformation (https://doi.org/10.1021/acs.analchem.2c03183). Our custom instrumentation and software (https://doi.org/10.1021/acs.analchem.1c05339) enable measurement of the amide hydrogen exchange rates of wild-type alpha-synuclein at amino acid resolution under physiological conditions, mimicking those in the extracellular, intracellular, and lysosomal compartments of cells (https://www.jove.com/t/64050/millisecond-hydrogendeuterium-exchange-mass-spectrometry-for-study). We now have the exciting possibility to experimentally measure sub-populations of conformers of intrinsically disordered proteins under physiological conditions, use this to build data-driven atomistic models and correlate these with biological and pathological function (http://www.nature.com/articles/s41467-020-16564-3).
Non-equilibrium protein structural dynamics
Whilst it is becoming routinely possible to determine accurate structural models of proteins with high resolution, it is still challenging to ascertain the specific structurally dynamic changes that underpin protein functional switching. The archetypal allosteric enzyme, glycogen phosphorylase (GlyP) is one of the most studied and has a substantial therapeutic potential in treating metabolic diseases and cancers. However, a lack of understanding of its complex regulation, mediated by dynamic structural changes, hinder its exploitation as a drug target. Here, we precisely locate dynamic structural changes upon allosteric activation of GlyP, by developing a time-resolved non-equilibrium millisecond hydrogen/deuterium-exchange mass spectrometry (HDX-MS) approach. We resolved obligate transient changes in localized structure that are absent when directly comparing active/inactive states of the enzyme, thus rationalizing the mechanism of action of an allosteric activator.
This approach has broad application to determine the structural kinetic mechanisms by which proteins are regulated. We are actively developing this approach to understand fundamental metabolic regulation, signal transduction and in quantum biology to understand the molecular basis for magnetosensation in migratory birds.
From our new understanding of how proteins move in response to – and to direct – their environment, we are interested to control them ourselves in order to create medicines, biosensors and other biotechnologies.
We revealed the specific structural dynamic pathway for activation of a designed biosensor that bioluminesces in response to binding a drug (https://doi.org/10.1038/s41467-022-28425-2).
We have exploited these principles of equilibrium and non-equilibrium protein structural dynamics to uncover the rules for successful design and engineering of switchable antibody drugs, which are dosed as an inactive pro-drug and then activate passively at the site of a tumour, for example (https://doi.org/10.1080/19420862.2022.2095701).
Cutting edge experimental and theoretical approaches
We are active in developing new instrumentation for millisecond time-resolved hydrogen/deuterium-exchange mass spectrometry, new mass spectrometry methods (http://pubs.acs.org/doi/abs/10.1021/acs.analchem.6b04158), new analysis software (https://doi.org/10.1021/acs.analchem.1c05339), new mathematical and statistical models (http://dx.doi.org/10.1016/j.jmb.2016.02.022). This agile interdisciplinary approach enables us to ask – and answer – the most challenging and exciting questions.
PhD Studentship Opportunity
Models of protein fate from models of cell fate
Dr JJ Phillips & Dr Kyle Wedgwood
Living Systems Institute, University of Exeter, UK
- Neurons are able to switch between functional states to transmit signals between cells.
- Protein molecules within the neuron, such as alpha-synuclein and glycogen phosphorylase, similarly switch between functional states to determine different cellular behaviours and respond to them.
- This enables important cellular functions (e.g. release of synaptic vesicles), but also can be the basis of pathophysiological function (e.g. aggregation, leading to neuronal necrotic cell death and, ultimately, Parkinson’s disease).
- Models of neuronal behaviour can be built to understand and predict the transient behaviour that is crucial to determining the information content in their signals.
- Using data on the potential energy and functional properties of start and end states, we will develop ODE-based mathematical models of the non-equilibrium neuronal protein dynamics, specifically alpha-synuclein and glycogen phosphorylase.
- Using the mathematical model, we will enumerate the possible transition pathways between functional states and predict which intermediate (non-observed) states orchestrate the transient dynamics.
- From these predictions, we will determine experimentally-realisable perturbations to the environment that have the potential to direct protein trajectories onto secondary or parallel pathways that lead to pathogenic states (e.g., involving aggregation).
- We are uniquely able to measure the structural dynamics of intrinsically disordered protein (such as alpha-synuclein or the active site gate in glycogen phosphorylase) under physiological conditions at high spatiotemporal resolution – per amino acid per millisecond.
- Using our bespoke experimental system, we will test our predictions on which perturbations lead to secondary protein dynamics pathways and hence establish a framework for uncovering ‘hidden’ pathways involving the non-observed states.
- Data from these experiments will used to further refine the mathematical model and to generate new predictions on secondary pathways.
- We will also apply approaches such as Sparse Identification of Nonlinear Dynamics (SINDy) to build robust mathematical models from the noisy mass spectrometry data.
- Ultimately, we will use this interdisciplinary approach to identify how the dynamic behaviour of individual and whole populations of neuronal proteins underpins neuronal cell function and pathophysiology.
- Enquiries welcome: firstname.lastname@example.org and K.C.A.Wedgwood@exeter.ac.uk
- Application deadline: 09 January 2023
- Apply here: https://www.exeter.ac.uk/study/funding/award/?id=4533Application