Structural Bioinformatics of Human Anoctamin ANO6–ANO10 and Their Water-Soluble QTY Analogs

Human integral membrane proteins of the anoctamin (ANO) family (ANO6–ANO10) function as ion channels and lipid scramblases in a wide range of biological processes, making them very important therapeutic targets. However, because these proteins contain highly hydrophobic transmembrane domains, they typically require detergent extraction and subsequent work, which complicates their expression and structural analysis. In this study, we applied the simple QTY code to design water-soluble analogs of five ANO family members (ANO6–ANO10) by replacing the hydrophobic amino acids leucine (L), isoleucine (I)/valine (V), and phenylalanine (F) with the hydrophilic residues glutamine (Q), threonine (T), and tyrosine (Y), respectively. The structures of these QTY analogs were predicted using AlphaFold 3 and compared with both experimentally determined cryo-EM structures and AlphaFold 3-predicted structures of the native proteins. Structural superposition produced low root mean square deviation (RMSD) values ranging from 0.293[Formula: see text]Å to 0.811[Formula: see text]Å, indicating that the overall alpha-helical structure was preserved well after QTY substitution. Furthermore, surface hydrophobic patches on the QTY analogs were significantly reduced. Most importantly, the internal geometry of the calcium-binding pocket was preserved well, suggesting its biological functionality. Our structural bioinformatics studies using the QTY code and AlphaFold 3 not only open new avenues for designing water-soluble membrane proteins but also enable the use of these QTY analogs as stable antigens for therapeutic monoclonal antibody discovery to specifically target the ANO family.