Project Summary/Abstract This proposal combines work on viral ion channels and de novo protein design. Our work on the M2 proton channel from influenza A virus focuses on the mechanism of inhibition and proton movement through the channel. M2 is also the target of the amantadine class of influenza drugs, and most isolates of influenza A virus are now amantadine-resistant. Crystallography, computation and 2DIR will be used to interrogate the mechanism and aid in design of drugs that inhibit M2 in amantadine-resistant strains of the virus. In parallel, we will expand our studies to examine the mechanism of conduction and inhibition of the E-proteins from coronaviruses, which have structures and functions largely similar to M2. De novo protein design provides a means to test and refine our understanding of protein structure and function. We are developing computational methods to design proteins that bind small molecules such as drugs and metalloporphyrins. We are also designing membrane proteins to elucidate the principles by which they fold and function. To probe the mechanisms of highly selective proton conduction proteins, we are designing proton-selective channels that test a hypothesis that networks of water molecules (water wires) can form dynamically and transiently through apolar stretches of the proton conduction pathway in proteins. The water wires would allow conduction of protons but not large hydrated alkali metal ions such as K+ or Na+. We also are engaged in design of proteins that bind to and transport nutrients such as amino acids across membranes, and devising screens to test them in vivo.