Johnson, Neil A.2; Liu, F2; Weeks, P. D.2; Hentzen, A. E.2; Kruse, H. P.2; Parker, J. J.2; Laursen, Mette5; Nissen, Poul6; Costa, C. J.2; Gatto, Craig2
1 Department of Molecular Biology, Faculty of Science, Aarhus University, Aarhus University2 Illinois State University in Normal3 DANDRITE - Nissen Group, DANDRITE, Interfaculty, Aarhus University4 Department of Molecular Biology and Genetics - DANDRITE, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University5 DANDRITE - Nissen Group, DANDRITE, Interfaculty, Aarhus University6 Department of Molecular Biology and Genetics - DANDRITE, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University
Recombinant Ca(2+)-ATPase from tomato (i.e. LCA1 for Lycopersicon esculentum [Since the identification and naming of LCA1, the scientific name for the tomato has been changed to Solanum lycopersicum.] Ca-ATPase) was heterologously expressed in yeast for structure-function characterization. We investigate the differences between plant and animal Ca pumps utilizing comparisons between chicken and rabbit SERCA-type pumps with Arabidopsis (ECA1) and tomato plant (LCA1) Ca(2+)-ATPases. Enzyme function was confirmed by the ability of each Ca(2+)-ATPase to rescue K616 growth on EGTA-containing agar and directly via in vitro ATP hydrolysis. We found LCA1 to be approximately 300-fold less sensitive to thapsigargin than animal SERCAs, whereas ECA1 was thapsigargin-resistant. LCA1 showed typical pharmacological sensitivities to cyclopiazonic acid, vanadate, and eosin, consistent with it being a P(IIA)-type Ca(2+)-ATPase. Possible amino acid changes responsible for the reduced plant thapsigargin-sensitivity are discussed. We found that LCA1 also complemented K616 yeast growth in the presence of Mn(2+), consistent with moving Mn(2+) into the secretory pathway and functionally compensating for the lack of secretory pathway Ca-ATPases (SPCAs) in plants.
Archives of Biochemistry and Biophysics, 2008, Vol 481, Issue 2, p. 157-168