Delimitation of the Pore in Tweety Homolog 1 Channels: A Model-guided Approach

Tweety homolog proteins have been indicated as ´putative anion channels´, but yet there is some lack of definitive data in this regard. Here, we focused on the human Tweety homolog 1 protein (hTTYH1), and elaborated a model based on residue-coevolution analysis and ab initio principles. This much satisfactory model, was tested through sited-directed mutagenesis of residues, R371 and F394. Mutation of these residues resulted in alterations in ion selectivity and blocker sensitivity, respectively, confirming the predictions from the model. Hence, our results strengthen the idea that these proteins are in fact anion channels. As well, our model can be used to propose further structure-function relationships. Similar approaches can be employed for other channels, which have unknown structure, due to the very technically demanding and sometimes uncertain process of crystallization of membrane proteins for X-ray crystallography.