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Ca2+ transport activities of inside-out vesicles prepared from density-separated erythrocytes from rat and human

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Abstract

The plasma membrane Ca2+ ATPase in erythrocytes is vital for the maintenance of intracellular Ca2+ levels. Since the cytoplasmic Ca2+ concentration is elevated in older erythrocytes, the properties of the Ca2+ transport ATPase were examined during cell aging using inside-out vesicles (IOVs) prepared from density-separated, young (less dense, Ey) and old (more dense, Eo) rat and human erythrocytes. The transport of Ca2+ and the coupled hydrolysis of ATP were measured using radiolabeled substrates. The calmodulin-independent Ca2+ transport activity (Ey, 38.8 vs. Eo, 23.3 nmols/min/mg IOV protein) and the Ca2+ dependent ATP phosphohydrolase activity (Ey, 53.5 vs. Eo, 48.8 nmols/min/mg protein) were greater in IOVs prepared from younger (less dense) rat erythrocytes. The calmodulin-independent Ca2+ transport activity in IOVs from human erythrocytes was 12.9 nmols/min/mg IOV protein for Ey and 10.7 nmols/min/mg IOV protein for Eo. Inside-out vesicles from older (more dense) cells exhibited a lower pumping efficiency as determined by the calculated stoichiometry, molecule of Ca2+ transported per molecule of ATP hydrolyzed (rat: Ey, 0.74 vs. Eo, 0.49; human: Ey, 1.22 vs. Eo, 0.77). The enzymatic activity of rat and human Ey IOVs was labile. The Ca2+ transport activity in Ey but not Eo IOVs rapidly declined during cold storage (4°C). The decrease in Ca2+ transport activity during aging may accentuate the age-related decline in several erythrocytic properties.

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Abbreviations

IOV:

Inside-Out Vesicles

Ey:

Erythrocytes enriched with young (less dense) cells

Eo:

Erythrocytes enriched with old (more dense) cells

ACEase:

Acetylcholinesterase

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  1. Norbert W. Seidler

    Present address: Department of Biochemistry & Molecular Biology, SUNY-Health Science Center at Syracuse, 766 Irving Avenue, 13210, Syracuse, NY, USA

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  1. Department of Biochemistry & Molecular Biology, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, and Graduate School of Biochemical Sciences, 07103, Newark, New Jersey, USA

    Norbert W. Seidler & Norbert I. Swislocki

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Seidler, N.W., Swislocki, N.I. Ca2+ transport activities of inside-out vesicles prepared from density-separated erythrocytes from rat and human. Mol Cell Biochem 105, 159–169 (1991). https://doi.org/10.1007/BF00227755

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  • DOI: https://doi.org/10.1007/BF00227755

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