Applications of Mass Spectrometry to Membrane Protein Drug Development, to address urgent work on COVID-19

Impact/national response: our aim is to accelerate effective drug development and provide critical data rapidly for continued research. We will determine novel chemical matter for further development, identify existing drugs for repurposing and highlight off-target drug binding Urgency: The type of data we are able to generate is needed now to feed into drug discovery programmes; it cannot be obtained through other experimental platforms in the same timescale (6 months)Our approach: we will probe associations in viral assembly and cellular entry pathways using our bespoke massspectrometry (MS) platform (Nature Methods in press). We will monitor the oligomeric status of proteins their ligand binding capacity and PTM status within individual experiments.Objective 1: inhibit Mpro through disruption of the monomer dimer interface.Mpro, a key CoV enzyme, mediates viral replication and transcription. Mpro shares considerable homology withMERS-CoV Mpro which showed substrate-induced dimerization with proteolytic activity. We have proof-ofconceptdata that show the monomer-dimer equilibrium enabling us to screen to disrupt dimer interactionsusing our unique MS platform.Objective 2: understand antigen processing and interactionsPrevious SARS-CoV infections suggest that glycosylation of a specific residue abrogates S protein binding implyingthat interfering with glycosylation could impede recognition. We will investigate how S protein glycans affectassociations with ACE2 to inform design of glycoprotein processing inhibitors.Objective 3: characterise the oligomeric states and binding sites of ACE2The SLC B0AT1 supports dimerization of ACE2. How do lipids, proteases (TMPRSS2) and other small moleculesregulate its activity? We will apply MS-based methods, optimized to study membrane proteins, protein-protein,and protein-ligand interactions to this complex.Feasibility/Critical mass: All proteins are available immediately through collaboration with the SGC Oxford. Wealso have unique chemical libraries that can be deployed for this project as well as bespoke instrumentation. Ourdepartment is open for COVID-19 work only. I have secured permission for access and have engaged a dedicatedteam of highly experienced researchers.Accessibility: High-throughput screens using X-ray structures are ongoing in Oxford and feed into the COVID-19Therapeutics Accelerator Consortium. We will communicate all our findings immediately to this, and other localand national initiatives, including national repositories where they can be accessed immediately. Wherepertinent, these (and other) findings will also be communicated to decision makers through my role as Presidentof the Royal Society of Chemistry.