Award details

A reverse chemical genetics and chemical biology approach to probing inositol 145-trisphosphate receptor function

Reference	BB/C515255/1
Principal Investigator / Supervisor	Professor Stuart Conway
Co-Investigators / Co-Supervisors	Professor Garry Taylor
Institution	University of St Andrews
Department	Chemistry
Funding type	Research
Value (£)	285,287
Status	Completed
Type	Research Grant
Start date	01/02/2005
End date	31/08/2008
Duration	43 months

Abstract

Ca2+ is an almost universal intracellular messenger, controlling a diverse range of cellular processes, such as gene transcription, muscle contraction and cell proliferation. In most cells Ca2+ has its major signalling function when its concentration is elevated in the cytosolic compartment. Berridge and Irvine were the first to demonstrate that activations of InsP3Rs causes an increase in intracellular Ca2+ concentrations. This occurs as a result of cell-surface receptor activation, which triggers the phospholipase C (PLC)-mediated hydrolysis of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) to form the second messengers, diacylglycerol (DAG) and InsP3. InsP3, which is hydrophilic, diffuses in to the cytosol and activates InsP3Rs. These tetrameric receptors are ligand-gated ion channels located within the endoplasmic reticulum (ER) membrane. Binding of InsP3 to InsP3Rs causes the channel to open releasing Ca2+ into the cytosol from a distinct store within the ER. Although there are a number of compounds that selectively activate InsP3Rs over RYRs and NAADPRs, there are currently no potent, selective InsP3R antagonists available. The existing antagonists of InsP3 action are not applicable in many situations due to poor specificity or lack of membrane permeability. The study of Ca2+ signalling and InsP3R pharmacology is of undoubted importance due to its ubiquitous nature within cellular processes and hence its involvement in cellular dysfunction. However, the lack of selective pharmacological tools for the InsP3Rs is hampering research into the biological roles of these important Ca2+ releasing channels. This project seeks to study the InsP3Rs by addressing the lack of selective, potent InsP3R antagonists. Using a multidisciplinary approach involving synthetic chemistry, molecular modelling, biophysical techniques and molecular biology, we will synthesise a range of enantiomerically pure compounds postulated to act as selective InsP3R antagonists. We will also express the InsP3 binding domains of InsP3Rs and using isothermal titration calorimetry (ITC), measure the thermodynamics of ligand binding. By labelling two specific surface cysteines of the binding domains with nitroxides we will be able to study the binding characteristics of InsP3 and the antagonists using electron spin coupling (ESR). Simultaneously, we will develop molecular models of the InsP3R isoforms. The binding of compounds that show InsP3R antagonist activity will be rationalised using molecular modelling. The results of this study will be employed in designing a second generation of compounds with either enhanced antagonists activity and or InsP3R isoform selectivity. Joint with BB/C515298/1

Summary

unavailable

Committee	Closed Committee - Biomolecular Sciences (BMS)
Research Topics	Structural Biology
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

Associated awards:

BB/C515298/1 A reverse chemical genetics and chemical biology approach to probing inostiol 145-trisphosphate receptor function

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