Structural studies of the Sulphonylurea receptor subunit of the ATP-sensitive potassium channel by electron crystallography and molecular modelling. (360G-Wellcome-080717_Z_06_Z)

Structural studies of the Sulphonylurea receptor subunit of the ATP-sensitive potassium channel by electron crystallography and molecular modelling To understand the relationship between KATP channel structure and function, for example how mutations cause neonatal diabetes and where nucleotides and drugs bind, we need structural information of the individual subunit proteins and the way these subunits interact within the KATP channel complex. This project aims to provide that information in two ways: by molecular modelling and simulation of the SUR1 subunits and their interaction with Kir6.2, and by determining the structure of SUR1 in two-dimensional crystals by electron crystallography. Determining structures of large eukaryotic membrane proteins to even near-atomic resolution is notoriously difficult. However, our current studies indicate that the electron crystallography approach is feasible and should result in a SUR1 structure at sub-10Å or better resolution within three years. Consequently, this proposal is structured into three parts which can be addressed independently but have obvious beneficial synergies. We expect that the proposed combination of different threads such as (i) cryo-electron microscopy, (ii) models based on existing ABC transporter structures, using established and novel modelling techniques, and (iii) a structural reinterpretation of existing functional data will lead to a better understanding of SUR1 and KATP channel structure over the course of this PhD project. Part 1: Two-dimensional crystallisation of purified SUR1 and SUR2 protein and method development in the image processing of micrographs of two-dimensional crystals. Part 2: Homology modelling and molecular dynamics simulations of SUR and the KATP channel. Part 3: Developing systematic methods for the interpretation of medium resolution structural data of membrane proteins.