Hyperactivation of the human plasma membrane Ca2+ pump PMCA h4xb by mutation of Glu99 to Lys
The transport of calcium to the extracellular space carried out by plasma membrane Ca2+ pumps (PMCAs) is essential for maintaining low Ca 2+ concentrations in the cytosol of eukaryotic cells. The activity of PMC Asis controlled by autoinhibition. Autoinhibition is relieved by the binding of Ca2+-cal...
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| Autores principales: | , |
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| Formato: | JOUR |
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| Acceso en línea: | http://hdl.handle.net/20.500.12110/paper_00219258_v289_n15_p10761_Mazzitelli |
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| Sumario: | The transport of calcium to the extracellular space carried out by plasma membrane Ca2+ pumps (PMCAs) is essential for maintaining low Ca 2+ concentrations in the cytosol of eukaryotic cells. The activity of PMC Asis controlled by autoinhibition. Autoinhibition is relieved by the binding of Ca2+-calmodulin to the calmodulin-binding autoinhibitory sequence, which in the human PMCA is located in the C-terminal segment and results in a PMCA of high maximal velocity of transport and high affinity for Ca2+. Autoinhibition involves the intramolecular interaction between the autoinhibitory domain and a not well defined region of the molecule near the catalytic site. Here we show that the fusion of GFPto theCterminus of the h4xbPMCA causes partial loss of autoinhibition by specifically increasing the Vmax. Mutation of residue Glu99 to Lys in the cytosolic portion of the M1 transmembrane helix at the other end of the molecule brought the Vmax of the h4xbPMCAto near that of the calmodulin-activated enzyme without increasing the apparent affinity for Ca2+. Altogether, the results suggest that the autoinhibitory interaction of the extreme C-terminal segment of the h4PMCAis disturbed by changes of negatively charged residues of the N-terminal region. This would be consistent with a recently proposed model of an autoinhibited form of the plant ACA8 pump, although some differences are noted. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc. |
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