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Understanding the structural basis of sodium-triggered activation of neuronal potassium channels

Reference	BB/X007251/1
Principal Investigator / Supervisor	Dr Jonathan Lippiat
Co-Investigators / Co-Supervisors	Dr Antreas Kalli, Dr Stephen Muench
Institution	University of Leeds
Department	Sch of Biomedical Sciences
Funding type	Research
Value (£)	526,905
Status	Current
Type	Research Grant
Start date	01/04/2023
End date	31/03/2026
Duration	36 months

Abstract

Sodium and voltage-activated potassium channels KNa1.1 and KNa1.2 ("KNa channels"), are two of the most poorly understood ion channels in the mammalian central and peripheral nervous systems. From studies of mice and humans, their normal functioning is required for diverse neurological functions such as cognition, neurodevelopment, intellect, motor learning, hearing and nociception. Of scientific importance is the understanding of the molecular basis of how these channels bind and respond to intracellular sodium and how transmembrane potential is sensed. Structural data indicate that these potassium channels possess neither any known sodium binding site nor voltage sensing S1-S4 transmembrane segments that are characteristic of related Kv channel subtypes. Available pharmacological tools, both inhibitors (e.g. bepridil, quinidine) and activators (e.g. loxapine, niclosamide), possess poor selectivity and weak potency. As such, caution is attached to their use in tissue or animal studies, or as starting points for compound discovery and development. Our pilot studies describe a structural change in the intracellular domains that is responsible for sodium activation and propose that voltage-dependent gating is controlled by the selectivity filter. We will build on this using a combination of molecular dynamics simulations, mutagenesis, electrophysiology, membrane protein structural analysis, and computational chemistry. We will describe the structural basis of KNa channel function and identify novel modulators that target domains unique to these channels. Our main objectives are: 1. To identify the mechanism of activation by sodium ions in the intracellular RCK2 domains 2. To identify the structural changes underlying gating at the selectivity filter 3. To describe how sodium sensing by RCK2 domains is coupled to activation of the selectivity filter gate 4. To identify chemical modulators that target regulatory sites to activate or inhichannel function

Summary

Similar to modern computers, the function of nerves throughout our brain and body requires tiny components to regulate charge movement and changes in voltage at exactly the right time and place. Unlike computers, however, the electrical activity in our body also responds to the chemical and physical environment due to its organic nature. The tiny components that control electrical activity in nerves are called ion channels and they control the movement of charged particles in the form of potassium, sodium, chloride, and calcium ions across the cell membrane. Understanding how they work and how they contribute to the normal functioning of the human body is of both scientific importance and also for the development of pharmacological tools that can fine-tune their activity. The protein of interest in this research proposal are potassium channels that respond primarily to the presence of sodium ions inside cells and secondarily the membrane voltage. These potassium channels are required for our brains and bodies to develop normally, and to understand and feel the world around us. Over the last forty years, laboratory techniques have existed that enable the charge flow across cell membranes through ion channels to be recorded from individual cells and even individual ion channel molecules. This has enabled researcher to describe how ion channels behave from the functional point of view. What researchers lack, however, is a description of what the proteins look like and how their structures change from one moment to the next in response to chemical and physical triggers. There have been some exciting developments in microscopy that enable static images of protein structures to be obtained. Similarly, computational tools have also been developed to allow us to show how these protein molecules change shape under certain conditions. Our research proposal aims to bring these state-of-the-art techniques together to understand the molecular basis of how these potassium channels work and how they respond to the presence of sodium ions and changes in membrane voltage. In carrying out this research, we will identify parts of the protein structure that could be targeted by chemicals to fine-tune the protein behaviour and will use computational tools to predict which chemicals may work. In doing so we will identify chemicals that either increase or decrease potassium channel function that could be used in further experiments to better understand the roles played by these proteins throughout the body. These may also be starting points for developing the pharmacology and therapeutics for diverse human conditions across the lifespan.

Committee	Research Committee D (Molecules, cells and industrial biotechnology)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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