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D. Tang, W. Dean, D. Borchman, C.A. Paterson; The Influence of Membrane Lipid Composition on Plasma Membrane Ca–ATPase Function . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1861.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Lens membrane lipid composition and Ca–ATPase activity both change with age, cataract and location in the lens. We explored relationships between lipid characteristics (composition and structure) and plasma membrane Ca–ATPase (PMCA) activity. Methods: PMCA isoform 4, one of the major isoforms of the PMCA in the human lens, was purified from human erythrocytes using a calmodulin affinity Sepharose column. PMCA4 was reconstituted into liposomes with a protocol that uses Triton X–100. PMCA activity was determined using a coupled enzyme assay. Fluorescence techniques were used to detect lipid–PMCA interactions and lipid dynamics. Results: Reconstitution into lipids was essential for PMCA4 activity. The activity was about 2.5 µmol/mg protein/min for purified PMCA4 reconstituted into lipids from human lens fiber cells or epithelial cells. There was no difference between the lipid dynamics of human lens fiber cells or epithelial cells. When purified PMCA4 was reconstituted into membrane vesicles composed of phosphatidylcholine, PMCA4 activity was 2 to three times higher when the acyl chains were ordered (stiff) compared to disordered (fluid) acyl chains. In a completely ordered lipid hydrocarbon chain environment, PMCA4 associates more strongly with the acidic lipid phosphatidylserine in comparison to phosphatidylcholine. PMCA4 associates much more strongly with phosphatidylcholine containing ordered hydrocarbon chains than disordered hydrocarbon chains. Conclusions: PMCA activity is influenced by membrane lipid composition and structure. The high degree of lipid order in human lens membranes may serve to support PMCA activity. The absence of PMCA activity in the cortical region of human lenses is apparently not due to a different lipid environment. The known rise in lipid order with age could account for the increase in PMCA activity with age.
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