The MPL was established to assist researchers investigating proteins that are embedded in the membranes that coat thousands of cells in the body. These proteins make up around 30% of the human genome. We have over 7,000 membrane proteins in our bodies and many of these are important drug targets; over 50% of current commercially available drugs target membrane proteins.
Crystallising membrane proteins is an essential, and extremely difficult step towards solving their structure. Having a dedicated laboratory with cutting edge equipment, close to the experimental stations where membrane protein structures can be solved, greatly enhances scientists’ ability to successfully crystallise membrane proteins and further our understanding of these important drug targets.
The MPL is open to user applications from anywhere in the world, and proteins crystallised here have been used in experiments in other facilities. A forthcoming paper will describe the successful XFEL data collection at SACLA-Japan from the MPL team, Diamond (I24 and XFEL team) and Oxford collaborators (Department of Biochemistry).
Recently published work1 details a collaboration between Brazilian scientists and the MPL, which investigated the role that two mitochondrial pyruvate carrier subunits, MPC1 and MPC2, play in the active transport of glycolytic pyruvate across the inner mitochondrial membrane. Diseases such as cancer, Alzheimer’s disease, and diabetes are known to be pyruvate-related, and a greater understanding of these membrane proteins will aid in the development of treatments for these diseases.
Research conducted elsewhere has proposed that MPC1 and MPC2 function together via the formation of an oligomeric structure. However, in this new work researchers provided an unprecedented in vitro demonstration that human MPC2 functions independently of MPC1 to induce pyruvate transport. They used synchrotron radiation circular dichroism (SRCD) analysis, on beamline B23, to investigate the secondary structure composition of human MPC2, and to gauge the conformational changes due to binding of substrate and inhibitor. The significant changes in the secondary structure content of MPC2 that they detected support the interaction between the protein and ligands.
These results are the first successful, large-scale, recombinant production and functional reconstitution of this family of solute carriers in an artificial lipid bilayer. It opens up a discussion concerning pyruvate import regulation by at least two different molecular entities in human mitochondria: heterotypic MPC1:MPC2 and homotypic MPC2:MPC2, and has immediate implications for the development of small-molecule-oriented therapeutics that specifically target MPC2 in pyruvate-related diseases.
Over the last decade, data collection and data processing landscape in X-ray crystallography for biomolecules has changed significantly. Improvements include faster readouts from silicon pixel detectors, ubiquitous presence of robotic arms and related cryogenics, new types of set-up for single and multiple crystal mounting, fast data transfer and larger capacity for data storage, new processing software and continued introduction and update of process pipelines. One consequence of these changes is the ability to create complete datasets from several crystals, rather than a single crystal.
Also in collaboration with the MPL research published in the journal Crystals2, discusses how the use of multiple crystals has become an accepted methodology in structural biology, with synchrotrons installing technology, hardware and software to make the technique routinely accessible to users. Diamond’s automated VMXi beamline is a prime example; handling large numbers of crystals for data collection is also the standard mode of operation for free-electrons laser sources. The article describes how to use the computer program BLEND to help assemble complete datasets for the solution of macromolecular structures derived from data collection from multiple crystals.
- Nagampalli RSK et al. doi:10.1038/s41598-018-21740-z
- Mylona A et al. doi:10.3390/cryst7080242