The activating mechanism of regucalcin, a calcium-binding protein isolated from rat liver cytosol, on (Ca2+−Mg2+)-ATPase in the plasma membranes of rat liver was investigated. (Ca2+−Mg2+)-ATPase activity was markedly increased by a sulfhydryl (SH) group protecting reagent dithiothreitol (DTT; 2.5 and 5 mM as a final concentration), while the enzyme activity was significantly decreased by a SH group modifying reagent N-ethylmaleimide (NEM; 0.5–5 mM). The effect of DTT (5 mM) to increase the enzyme activity was clearly blocked by NEM (5 mM). Regucalcin (0.25–1.0 μM) significantly increased (Ca2+-Mg2+)-ATPase activity. This increase was completely blocked by NEM (5 mM). Meanwhile, digitonin (0.04%), which can solubilize the membranous lipids, significantly decreased (Ca2+−Mg2+)-ATPase activity. Digitonin did not have an effect on the DTT (5 mM)-increased enzyme activity. However, the effect of regucalcin (0.25 μM) increasing (Ca2+−Mg2+)-ATPase activity was entirely blocked by the presence of digitonin. The present results suggest that regucalcin activates (Ca2+−Mg2+)-ATPase by the binding to liver plasma membrane lipids, and that the activation is involved in the SH groups which are an active site of the enzyme.
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Rasmussen J: Cell communication, calcium ion, and cyclic adenosime monophosphate. Science 170: 404–412, 1970
Cheung WY: Calmodulin plays a pivotal role in cellular regulation. Science 202: 19–27, 1984
Williamson JR, Cooper RH, Hoek JB: Role of calcium in the hormonal regulation of liver metabolism. Biochim Biophys Acta 639: 243–295, 1981
Reinhart PH, Taylor WM, Bygrave FL: The role of calcium ions in the mechanisms of action of α-adrenergic agonists in rat liver. Biochem J 223: 1–13, 1984
Yamaguchi M, Yamamoto T: Purification of calcium binding substance from soluble fraction of normal rat liver. Chem Pharm Bull 26: 1915–1918, 1978
Shimokawa N, Yamaguchi M: Molecular cloning and sequencing of the cDNA coding from a calcium-binding protein regucalcin from rat liver. FEBS Lett 327: 251–255, 1993
Yamaguchi M, Mori S: Inhibitory effect of calcium-binding protein regucalcin on protein kinase C activity in rat liver cytosol. Biochem Med Metab Biol 43: 140–146, 1990
Yamaguchi M, Tai H: Inhibitory effect of calcium-binding protein regucalcin on Ca2+/calmodulin-dependent cyclic nucleotide phophodiesterase activity in rat liver cytosol. Mol Cell Biochem 106: 25–30, 1991
Yamaguchi M, Sakurai T: Inhibitory effect of calcium-binding protein regucalcin on Ca2+-activated DNA fragmentation in rat liver nuclei. FEBS Lett 279: 281–284, 1991
Yamaguchi M, Sakurai T: Reversible effect of calcium-binding protein regucalcin on Ca2+-induced inhibition of deoxyuridine 5′-triphosphatase activity in rat liver cytosol. Mol Cell Biochem 110: 25–29, 1992
Lotersztjn S, Hanoune J, Pecker F: A high affinity calcium-stimulated magnesium-dependentATPase in rat liver plasma membranes. Dependence on an endogenous protein activator distinct from calmodulin. J Biol Chem 256: 11209–11215, 1981
Chen K-M, Junger KD: Calcium transport and phosphorylated intermediate of (Ca2+-Mg2+)-ATPase in plasma membranes of rat liver. J Biol Chem 258: 4404–4410, 1983
Yamaguchi M, Mori S, Kato S: Calcium-binding protein regucalcin is an activator of (Ca2+−Mg2+)-adenosine triphosphatase in the plasma membranes of rat liver. Chem Pharm Bull 36: 3532–3539, 1988
Takahashi H, Yamaguchi M: Regulatory effect of regucalcin on (Ca2+−Mg2+)-ATPase in rat liver plasma membranes: comparison with the activiation by Mn2+ and Co2+. Mol Cell Biochem 124: 169–174, 1993
Takahasahi H, Yamaguchi M: Regucalcin modulates hormonal effect on (Ca2+−Mg2+)-ATPase activity in rat liver plasma membranes. Mol Cell Biochem 125: 171–177, 1993
Lowry OH, Rosebrough NH, Farr AL, Randall RF: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–273, 1951
Prpić V, Green KC, Blackmore PF, Exton JH: Vassopressin-, angiotensin II-, and α -adrenergic-induced inhibition of Ca2+ transport by rat liver plasma membrane vesicles. J Biol Chem 259: 1382–1385, 1984
Nakamura M, Mori K: Colorimetric determination of inorganic phosphorus in the presence of glucose-1-phosphate and adenosine triphosphate. Nature 182: 1441–1442, 1958
Cullen PJ, Comerford JG, Dawson AP: Heparin inhibits the inositol 1,4,5-triphosphate-induced Ca2+ release from rat liver microsomes. FEBS Lett 228: 57–59, 1988
Lin S-H, Wallace MA, Fain JN: Regulation of (Ca2+−Mg2+)-ATPase activity in hepatocyte plasma membranes by vasopressin and phenylephrine. Endocrinology 113: 2268–2275, 1983
Lotersztajn S, Epand RM, Mallat A, Pecker F: Inhibition by glucagon of the calcium pump in liver plasma membranes. J Biol Chem 259: 8195–8201, 1984
Pecker F, Lotersztajn S: Fe2+ and other divalent metal ions uncouple Ca2+ transport from (Ca2+−Mg2+)-ATPase in rat liver plasma membranes. J Biol Chem 260: 731–735, 1985
Murphy EK, Coll TL, Rich TL, Williamson JR: Hormonal effects on calcium homeostasis in isolated hepatocytes. J Biol Chem 255: 6600–6608, 1980
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Takahashi, H., Yamaguchi, M. Activating effect of regucalcin on (Ca2+-Mg2+)-ATPase in rat liver plasma membranes: relation to sulfhydryl group. Mol Cell Biochem 136, 71–76 (1994). https://doi.org/10.1007/BF00931607
- SH group
- rat liver plasma membrane