De novo protein design by citizen scientists

Online citizen science projects such as GalaxyZoo(1), Eyewire(2) and Phylo(3) have been very successful for data collection, annotation, and processing, but for the most part have harnessed human pattern recognition skills rather than human creativity. An exception is the game EteRNA(4), in which game players learn to build new RNA structures by exploring the discrete two-dimensional space of Watson-Crick base pairing possibilities. Building new proteins, however, is a more challenging task to present in a game, as both the representation and evaluation of a protein structure are intrinsically three-dimensional. We posed the challenge of de novo protein design in the online protein folding game Foldit(5). Players were presented with a fully extended peptide chain and challenged to craft a folded protein structure with an amino acid sequence encoding that structure. After many iterations of player design, analysis of the top scoring solutions, and subsequent game improvement, Foldit players can now, starting from an extended polypeptide chain, generate a diversity of protein structures and sequences which encode them in silico. 146 Foldit player designs with sequences unrelated to naturally occurring proteins were encoded in synthetic genes; 56 were found to be expressed in E. coli with good solubility and to adopt stable monomeric folded structures in solution. The diversity of these structures is unprecedented in de novo protein design, representing 20 different folds—including a new fold not observed in natural proteins. High resolution structures were determined for four of the designs, and are nearly identical to the player models. This work makes explicit the considerable implicit knowledge contributing to success in de novo protein design, and shows that citizen scientists can discover creative new solutions to outstanding scientific challenges, such as the protein design problem.