Single Molecular Receptor Dynamics

The transmission of nerve signals in the body and brain is dependent on proteins called receptors. Most neurotransmission that governs memory and learning is controlled by ionotropic glutamate receptors, so-called, because upon binding of glutamate (the neurotransmitter) they open a pore into the neuron that allows positively charged ions (sodium and potassium) to pass through it. This is the basis of all nerve signals in the brain. It is therefore perhaps unsurprising that glutamate receptors have been implicated in many neurological conditions of the central nervous system (CNS) ranging from epilepsy to Alzheimer's disease. Despite much progress, exactly how the receptor changes conformation (shape) when it binds glutamate sill remains unclear.

In our previous work, we were able to show, for one particular sub-family of receptors called kainate receptors, how a region away from the glutamate-binding site could control the dynamic properties of the receptor. In particular, this region contains separate binding sites for sodium and chloride ions. Our previous work demonstrated that a collaborative approach which combined our expertize in computational biochemistry with experimental expertize of colleagues in Canada was extremely powerful in understanding the way in which these receptors work.

In this proposal we want to apply what we have learnt on the kainate receptors to another major subfamily of ionotropic glutamate receptors, the AMPA receptors. The work focuses on understanding how the composition of a key interface within the receptor controls which conformations the receptor can access. The knowledge we gain here, can in the longer-term be very useful for the design of new compounds that target neurological diseases.

Our proposal utilizes the power of molecular simulations to provide atomic-level detail of what controls the way this binding site behaves. A full understanding of this is necessary if we are to have any chance of developing compounds that act in a predictable way. The results we will generate will be verified and tested by our collaborators at McGill University, Canada and also at the University of Copenhagen, Denmark.

Ionotropic glutamate receptors (iGluRs) mediate fast neurotransmission in the brain and central nervous system and consequently are linked with the processes of memory and learning. Previous research has shown that in one subtype of iGluRs, the kainate receptors (KARs), an allosteric site that binds cations and anions is directly associated with controlling the receptor moving from the open state into the desensitized state. Recently we also showed how the integrity of this site is controlled at the atomic level using molecular dynamics simulations in conjunction with single-channel experiments and site-directed mutagenesis. The significance of this work was that it enabled a different picture of the underlying mechanism to be formulated that is consistent and unifies both new and old data. For some aspects of that work, it became apparent that MD simulations were the only method that could provide a molecular interpretation of the data.

In this proposal, we will apply this approach to the AMPA receptors, which are the major class of receptors responsible for the transfer of a nerve impulse across a synapse. The work will exploit our expertise in molecular dynamics simulations including the calculation of free energies of binding (via thermodynamic integration) and potential of mean force (PMF) calculations to examine the stability of the ligand-binding domain interface and make experimentally testable predictions.

The work will be high impact and has consequences for how we understand the function of these receptors. Key to this proposal is the collaboration with the groups of Derek Bowie at McGill, where the single channel experiments will be performed and Jette Kastrup at the University of Copenhagen where X-ray crystallography will be performed. Those aspects are being funded independently, and thus this proposal, represents exceptional value for money for the BBSRC as the project will benefit directly from an already proven collaboration.

The work proposed here, which is aimed at understanding how ionotropic glutamate receptors work is very much at the level of basic research. Consequently, the impact when it arises, will take time to come to fruition. However, as we are utilizing our simulations to interpret single channel data in a collaborative fashion, the impact is potentially high as it can provide a different way to view the mechanism of the receptor. The beauty is that this approach can also account for other pre-existing data including crystallographic and macroscopic channel data.

There will ultimately be many different beneficiaries of the research outside of the academic circle. In the long-term, the most obvious group will be the general public and in particular patient groups who are suffering from neurological conditions. This group would specifically include sufferers of epilepsy and chronic pain, but by extension to other subtypes, would include other neurological conditions. Thus there is a direct impact on the nation's health. Our prior work has already produced important information about the dynamics of these receptors and that understanding is critical to us designing better and improved drugs in the future.

Aside from this group, the main beneficiary will be the pharmaceutical industry, which although currently going though a period of upheaval and rationalization, particularly for neuroscience programs, will still benefit from early basic-level research that puts any drug discovery process on a firmer footing. The drug-design process is still extremely difficult and time-consuming. Drugs are developed at vast expense, typically by screening large numbers of compounds and using large numbers of experimental animals. Only a handful of drugs have been discovered by designing them to fit a particular protein target. For channel proteins like ionotropic glutamate receptors, there are two key stumbling blocks: The issue of side-effects - ie making the compounds selective enough that we don't "hit" receptors that are functioning normally and secondly that actually we don't understand channel function well enough, especially with regards to how they change in shape when they are functioning in the body, regardless of disease state. It will take a long time to get to design drugs mostly in silico, but our approach outlined here have the demonstrated power to take this forward in the right direction. The UK has a big tradition and massive knowledge in drug discovery, partly because much of the basic science discoveries occurred here.

Thus, the basic research we are proposing here will, in the long term, benefit our society through its impact both on human health and well-being (drug discovery, better understanding of physiological and pathological processes) and on economic productivity (development of novel drugs). These processes will take a long time - the drug discovery process is usually between 10 and 20 years, but the impact can be long lasting and life-changing.