High-Throughput Screening for Ligands of the HEPN Domain of Sacsin Xinlu Li Marie Ménade Guennadi Kozlov Zheping Hu Zheng Dai Peter S. McPherson Bernard Brais Kalle Gehring 10.1371/journal.pone.0137298 https://plos.figshare.com/articles/dataset/_High_Throughput_Screening_for_Ligands_of_the_HEPN_Domain_of_Sacsin_/1541287 <div><p>Sacsin is a large protein implicated in the neurodevelopmental and neurodegenerative disease autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS), which features the loss of Purkinje neurons in the cerebellum. Although the domain architecture of sacsin suggests that it is a neuronal chaperone assisting in protein quality control, the precise function of sacsin remains elusive. Using fluorescence polarization (FP) assays, we confirmed that the HEPN domain of sacsin binds to nucleotides with low micromolar affinities. FP competition assays with a variety of nucleotides and nucleotide analogs revealed that the binding is primarily mediated by the phosphate groups of nucleotides. A high-throughput screen subsequently identified novel small molecule ligands of HEPN, providing new chemical probes for cell culture studies and drug development. Together, the results are consistent with the HEPN domain contributing to the functional activity of sacsin by binding to nucleotides or other multiply charged anionic compounds in neurons.</p></div> 2015-09-14 02:46:08 cell culture studies neuron binding neurodegenerative disease autosomal recessive spastic ataxia ARSACS sacsin protein quality control nucleotide HEPN domain FP competition assays