Time resolved structural studies are the new frontier in structure biology. In a time-resolved experiment we aim to directly visualise the conformational changes associated with biological macromolecular function. To date, no time-resolved structural data for an NSS family member exist, although we do have structures of putative intermediates along the transport pathway.
Project 10 aims to directly visualise in real-time, with high spatial resolution, the structural changes that couple substrate binding to transport of at least one of the NSS family members using time-resolved crystallography.
This requires determining the optimal sample delivery method for each system and developing a reaction initiation protocol, mostly likely using a photocaged substrate. In this project we will explore the possibilities offered by both X-ray and electron crystallography. For the latter, the ability to carry out diffraction studies under ambient conditions is important, and so a key aspect of the project will also be the development of suitable sample environments for room temperature electron diffraction experiments.
This project will be carried out in close collaboration with the rest of the network. The highly pure NSS family members that are generated in the other projects will be targets for microcrystallization to yield the samples needed for this project. The data we obtain in this project will be fed into the molecular modelling, in combination with the spectroscopic and biochemical data obtained in the other projects, to allow us to build a detailed molecular description of how this class of membrane transporters work.