Towards the classification of DYT6 dystonia mutants in the DNA-binding domain of THAP1

The transcription factor THAP1 (THanatos Associated Protein 1) has emerged recently as the cause of DYT6 primary dystonia, a type of rare, familial and mostly early-onset syndrome that leads to involuntary muscle contractions. Many of the mutations described in the DYT6 patients fall within the sequence-specific DNA-binding domain (THAP domain) of THAP1 and are believed to negatively affect DNA binding. Here, we have used an integrated approach combining spectroscopic (NMR, fluorescence, DSF) and calorimetric (ITC) methods to evaluate the effect of missense mutations, within the THAP domain, on the structure, stability and DNA binding. Our study demonstrates that none of the mutations investigated failed to bind DNA and some of them even bind DNA stronger than the wild-type protein. However, some mutations could alter DNA-binding specificity. Furthermore, the most striking effect is the decrease of stability observed for mutations at positions affecting the zinc coordination, the hydrophobic core or the C-terminal AVPTIF motif, with unfolding temperatures ranging from 46 degrees C for the wild-type to below 37 degrees C for two mutations. These findings suggest that reduction in population of folded protein under physiological conditions could also account for the disease.