Hepatic microsomal Ca2+-dependent ATPase. Calmodulin-dependence and partial purification

Moore, Pamela B.; Kraus‐Friedmann, Naomi

doi:10.1042/bj2140069

Cited by 25 publications

(9 citation statements)

References 25 publications

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“…45 Ca 2ϩ uptake was measured by the Millipore filtration technique [Moore and Kraus-Friedmann, 1983]. About 100-150 µg of protein/ml reaction of mixture was incubated for 1 min 37°C in 1 ml of medium containing 100 mM KCl, 20 mM Hepes, 1 mM NaN 3 , 1 mM MgCl 2 , 1 µM ruthenium red, and 100 µM CaCl 2 containing 45 Ca 2ϩ (0.185 MBq), pH 6.8.…”

Section: Atp-dependent 45 Ca 2؉ Uptakementioning

confidence: 99%

Role of regucalcin as an activator of Ca2+-ATPase activity in rat liver microsomes

Takahashi

Yamaguchi

1999

J. Cell. Biochem.

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The effect of Ca(2+)-binding protein regucalcin on Ca(2+)-ATPase activity in isolated rat liver microsomes was investigated. The presence of regucalcin (0.1-1.0 microM) in the enzyme reaction mixture led to a significant increase in Ca(2+)-ATPase activity. Regucalcin significantly stimulated ATP-dependent (45)Ca(2+) uptake by the microsomes. Thapsigargin (10(-6) M), a specific inhibitor of microsomal Ca(2+) pump enzyme (Ca(2+)-ATPase), clearly inhibited regucalcin (0.5 microM)-increased microsomal Ca(2+)-ATPase activity. Liver microsomal Ca(2+)-ATPase activity was markedly decreased by N-ethylmaleimide (NEM; 2.5 mM), while the activity was clearly elevated by dithiothreitol (DTT; 2.5 mM), indicating that the sulfhydryl (SH) group of the enzyme is an active site. The effect of regucalcin (0.5 microM) in increasing Ca(2+)-ATPase activity was completely inhibited by the presence of NEM (2.5 mM) or digitonin (10(-2) %), a solubilizing reagent of membranous lipids. Moreover, the effect of regucalcin on enzyme activity was seen in the presence of Ca(2+) ionophore (A23187; 10(-7) M). The present study demonstrates that regucalcin can stimulate Ca(2+) pump activity in rat liver microsomes, and that the protein may act the SH groups of microsomal Ca(2+)-ATPase.

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Section: Atp-dependent 45 Ca 2؉ Uptakementioning

confidence: 99%

Role of regucalcin as an activator of Ca2+-ATPase activity in rat liver microsomes

Takahashi

Yamaguchi

1999

J. Cell. Biochem.

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“…It has a high affinity for Ca2+ (K, 0.1 5-0.2 ,MM) and requires both Ca2+ and Mg2+-ATP for activity. T h e energy for Ca2' transport is derived from the hydrolysis of ATP and the Ca2+ transport process is inhibited by high concentrations of vanadate (Dawson & Fulton, 1983; Epping & Bygrave, 1984a, b; Schanne & Moore, 1986), but stimulated by oxalate (Dawson, 1982;Epping & Bygrave, 1984b), calmodulin (Moore & Kraus-Friedmann, 1983; but see Schutz & Soling, 1987) and glucose 6-phosphate (Benedetti et al, 1985(Benedetti et al, , 1986(Benedetti et al, , 1987. On the other hand, Ca2+ uptake by mitochondria occurs through the Ca2+ uniporter (K, for Ca2' uptake of 2-10 ,MM) in the inner mitochondrial membrane (Bygrave, I 978 a).…”

Section: (3) Intracellular Ca2+ Transportmentioning

confidence: 99%

Second Messengers and the Regulation of Ca 2+ Fluxes by Ca 2+ ‐mobilizing Agonists in Rat Liver

Altin

Bygrave

1988

Biological Reviews

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Summary Knowledge of the mechanism of action of Ca2+‐mobilizing agonists in liver has progressed considerably following the discovery that their interaction with specific receptors on the plasma membrane is accompanied by the hydrolysis of PIP2 and the generation of the second messengers diacylglycerol and IP3, for the activation of protein kinase C and the mobilization of intracellular Ca2+, respectively. Although the second messenger functions of diacylglycerol and IP3 in these actions seem well established, it is not yet clear how the agonists are able to regulate Ca2+ influx across the plasma membrane, an event which is crucial for those actions of the agonists which are dependent on the maintenance of an elevated level of cytosolic Ca2+, Whilst there is evidence for the existence of more than one pathway for Ca2+ influx in liver, it appears that in each instance the Ca2+ influx process is regulated differently to the Ca2+ influx through the volage‐sensitive Ca2+ channels that is known to occur in excitable tissues. At present it is not clear whether any of the Ca2+ influx pathways in liver is regulated by direct coupling to the agonist receptor mechanism on the outer surface of the plasma membrane, or whether the regulation involves the production of some second messenger(s). However, indirect evidence from a number of tissues appears to favour the involvement of both IP3 and IP4 in the regulation of Ca2+ influx. The mechanism by which IP3 and IP4 may regulate Ca2+ influx remains to be established, but it has been proposed that Ca2+ entry into the cell occurs through a pathway connecting the plasma membrane and the endoplasmic reticulum, following the release of intracellular Ca2+ from the lumen of the endoplasmic reticulum. Although it is not yet known whether glucagon (or cyclic AMP) activates the same pathway for Ca2+ influx as Ca2+‐mobilizing agonists, the marked potentiation by cyclic AMP of the Ca2+ influx induced by Ca2+‐mobilizing agonists has provided a powerful system with which to study the regulation of Ca2+ influx in liver. Whether this Ca2+ influx process occurs through some ion exchange mechanism (such as Ca2+/Na+ exchange) remains to be determined. Results from this study suggests that the Ca2+ influx is inhibited by neomycin, acidic pH, and a depolarization of the plasma membrane. The observation that cyclic AMP synergistically potentiates the influx of Ca2+ induced by Ca2+‐mobilizing agonists, that this influx appears to correlate with the reported ability of these agonists to induce PIP2 hydrolysis and accumulation of IP3, and that cyclic AMP synergistically potentiates the production of IP4 by vasopressin, are all consistent with the notion that IP3 and IP4 are involved in regulating Ca2+ influx. Whilst little is known about the Ca2+ transport process itself, these studies coupled with the recent finding that Ca2+ influx into the liver cell can occur through different pathways, seem set to lead to a better understanding of this important process in the near future.

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“…The microsomal fraction was prepared from the livers of male fed Sprague-Dawley rats as described by Moore & Kraus-Friedmann (1983). Excised livers were minced and homogenized in ice-cold 250mM-sucrose/lOmM-Hepes/0.2mM-CaC12, pH 7.2, with a motor-driven homogenizer fitted with a Teflon pestle.…”

Section: Microsomal Preparationmentioning

confidence: 99%

Phosphorylated intermediates of two hepatic microsomal ATPases

Fleschner¹,

Kraus‐Friedmann²,

Wibert³

1985

Biochemical Journal

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The hepatic microsomal Ca2+- and Mg2+-dependent ATPase phosphoenzyme intermediates were distinguished by using the chelators EGTA and CDTA (trans-cyclohexane-1,2-diamine-NNN'N'-tetra-acetic acid). The Ca2+-ATPase intermediate is a hydroxylamine-labile base-labile 125 000-Mr phosphoprotein. The Mg2+-ATPase intermediate is a hydroxylamine-stable base-stable 30 000-Mr phosphoprotein. This enzyme intermediate probably reflects the large basal ATPase activity of hepatic microsomal fraction. It is dependent on Mg2+, since formation of the phosphoenzyme is abolished in the presence of CDTA. Under these conditions, the basal ATPase activity is dramatically decreased. These data demonstrate two separate and distinct enzymes which are responsible for the two ATPase activities of hepatic microsomal fraction. Furthermore, these data indicate that more meaningful data about the microsomal Ca2+-ATPase might be obtained if the free ion concentrations are controlled with CDTA.

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Hepatic microsomal Ca2+-dependent ATPase. Calmodulin-dependence and partial purification

Cited by 25 publications

References 25 publications

Role of regucalcin as an activator of Ca2+-ATPase activity in rat liver microsomes

Role of regucalcin as an activator of Ca2+-ATPase activity in rat liver microsomes

Second Messengers and the Regulation of Ca 2+ Fluxes by Ca 2+ ‐mobilizing Agonists in Rat Liver

Phosphorylated intermediates of two hepatic microsomal ATPases

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