STRUCTURAL STUDIES OF SACSIN AND EDD UBIQUITIN LIGASE

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Autosomal recessive spastic ataxia of Charlevoix¿¿"Saguenay (SACS) is an early-onset neurodegenerative disease with high prevalence in the Charlevoix¿¿"Saguenay¿¿"Lac-Saint-Jean region of Quebec (Canada). The gene responsible for SACS codes for sacsin, a protein containing more than 4000 residues. In order to better understand the function of this protein and its relation to the disease, we initiated structural studies of individual domains of sacsin. We recently obtained crystals of HEPN and UBL domains of sacsin. Both crystals diffract to 2.8-2.9A at the home source, but no solution was found by molecular replacement using distantly related structures (below 30% identity). We plan to improve resolution of the diffraction data and find experimental phases using MAD experiments. The tumor suppressor hyperplastic discs protein (HYD), also known as EDD (E3 isolated by differential display) or UBR5, is a member of the family of HECT (homologous to E6-AP carboxyl terminus) E3 ubiquitin ligases, which target specific proteins for ubiquitin-mediated proteolysis. Mutations in EDD fail to properly terminate proliferation leading to tumors and also result in developmental abnormalities such as adult sterility due to germ cells defects. We crystallized HECT domain of EDD. The crystals diffract to 3.0A at the home source. No molecular replacement solution was found. We are going to use MAD experiment for experimental phasing and to improve resolution of the crystals. The structure will be important to study a substrate specificity of EDD.