Sub-cellular localization of two P-type ATPases, Atp13a1 and Atp13a2

CHED 922

Ryan Baldridge, baldridgery@nku.edu1, Adam C. Ketron, ketrona@nku.edu1, Jennifer Walter1, John Ledford1, Diana L. McGill, mcgill@nku.edu1, Patrick Schultheis1, and Kristi H. Martines, martines@nku.edu2. (1) Department of Chemistry, Northern Kentucky University, Nunn Dr, SC 204f, Highland Heights, KY 41099-1905, (2) Department of Biological Sciences, Northern Kentucky University, Nunn Dr, Highland Heights, KY 41099
P-type ATPases are membrane proteins that transport substrates against their concentration gradients. The focus of this study is two novel P-type ATPases: Atp13a1 and Atp13a2. To date, nothing is known about their substrate specificity, sub-cellular localization or function. The aim of this study was to determine the membrane location of Atp13a1 and Atp13a2 via western blotting and immunofluorescence. HeLa cell lines expressing GFP and V5-His tagged versions of Atp13a1 and Atp13a2 were produced. Lysates from these cell lines were layered onto a sucrose density gradient (40%-10% sucrose) and were centrifuged at 100,000g for 16 hours. One milliliter fractions were harvested from the gradient after centrifugation and tested via western blot. Antibodies to organelle markers in ER, Golgi and plasma membrane were used along with antibodies to the GFP and V5-His tags to co-localize the ATPases with the organelles. To verify the sub-cellular fractionation data, immunofluorescence was also used.
 

Undergraduate Research Poster Session: Biochemistry
2:00 PM-4:00 PM, Monday, March 26, 2007 Hyatt Regency Chicago -- Riverside Center, Poster

Division of Chemical Education

The 233rd ACS National Meeting, Chicago, IL, March 25-29, 2007