UC Berkeley

Ion channels are a large and diverse family of proteins broadly implicated in human physiology and that of other organisms. The study of the molecular mechanism of ion channels benefits from both “top-down” physiological and “bottom-up” biophysical studies. Here, we use a reductionist biophysical approach, with a focus on the use of cryo-electron microscopy (cryo-EM) to study the structure and function of three putative ion channels. The first, ORF3a is a putative ion channel encoded in the SARS-CoV-2 genome. Here, we determined the first ORF3a structure, and characterized ORF3a activity through electrophysiological recordings and fluorescent imaging of reconstituted ORF3a proteoliposomes. The second, Tweety-homolog family (TTYH) proteins, previously characterized as putative volume-regulated anion channels. Through parallel functional and structural studies, our work determined that TTYHs are not pore-forming subunits of anion channels. Additionally, we discovered a previously unknown “trans-dimeric” conformation of TTYH2. The third putative ion channel is TMEM87A, previously implicated in mechanosensitive conduction of cations. Here we determine the first structure of an TMEM87 protein, and through structural and bioinformatic analysis, identify related proteins forming the basis for a new protein family. Collectively, this work has demonstrated the value of cryo-EM for structural studies of small (<100 kDa) membrane proteins, while laying the foundation for deeper mechanistic understanding of each of these different proteins.