ERcalcistorin/protein-disulfide isomerase acts as a calcium storage protein in the endoplasmic reticulum of a living cell. Comparison with calreticulin and calsequestrin.
ERcalcistorin/protein-disulfide isomerase (ECaSt/PDI), a high capacity low affinity Ca2+-binding protein in the endoplasmic reticulum of sea urchin eggs (Lebeche, D., and Kaminer, B. (1992) Biochem. J. 287, 741-747), shares 55% sequence identity with mammalian PDI and has PDI activity (Lucero, H., Lebeche, D., and Kaminer, B. (1994) J. Biol. Chem. 269, 23112-23119). We report on ECaSt/PDI functioning as a Ca2+ storage protein in the endoplasmic reticulum (ER) of a living cell and compare it with calsequestrin and calreticulin, high capacity low affinity Ca2+-binding proteins in the sarcoplasmic reticulum and ER, respectively. Stably transfected Chinese hamster ovary cell clones expressed these proteins, which were localized in the ER of the cell. Microsomes from cells expressing ECaSt/PDI, calreticulin, and calsequestrin accumulated 17.2 +/- 0.27, 20.0 +/- 0.82, and 38.0 +/- 0.28 nmol of Ca2+/mg of protein, respectively; control microsomes accumulated from 2.6 +/- 0.17 to 2.9 +/- 0.14 nmol of Ca2+/mg of protein. The initial rate of Ca2+ uptake was similar in microsomes from transfected and control cells. Microsomes containing an ECaSt/PDI mutant in which 45% of the acidic residue pairs in the C terminus were truncated had a reduced Ca2+ storage capacity. This supports our previous hypothesis that the degree of low affinity Ca2+ binding is dependent on the number of pairs of carboxyl groups in the molecule. The maximal Ca2+ accumulation by microsomes containing the expressed ECaSt/PDI, C-terminally truncated ECaSt/PDI, calreticulin, or calsequestrin correlates approximately with the Ca2+ binding capacity of the respective proteins.
Lucero
H A
HA
Department of Physiology, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Lebeche
D
D
Kaminer
B
B
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis / Q000378:metabolism*; D037282:Calreticulin; D002155:Calsequestrin / Q000378:metabolism*; D006224:Cricetinae; D004527:Egg Proteins; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D007700:Kinetics; D008854:Microscopy, Electron; D008861:Microsomes / Q000378:metabolism* / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D014162:Transfection
D002135:Calcium-Binding Proteins; D037282:Calreticulin; D002155:Calsequestrin; C089323:ERcalcistorin protein, Strongylocentrotus purpuratus; D004527:Egg Proteins; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D019704:Protein Disulfide-Isomerases; D002118:Calcium