The sarcoplasmic reticulum Ca2+-ATPase

Møller, Jesper; Andersen, Johannes Lund; Maire, Marc le

doi:10.1007/bf00222696

Cited by 113 publications

(35 citation statements)

References 185 publications

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“…61), the ElP-E2P transition rate was almost as low as for membrane-bound Ca-ATPase and no decomposition of phosphoenzyme occurred. Deoxycholate is known to be a very efficient detergent for solubilization of Ca-ATPase in monomeric form, even at high protein concentration [27]. In the present study we observed a nearly homogenous sedimenting boundary (more than 90% sedimenting with S20,w = 4.8 S corresponding to monomer) when the sample used for the experiments of Fig.…”

Section: Discussionsupporting

confidence: 67%

Effect of phospholipid, detergent and protein‐protein interaction on stability and phosphoenzyme isomerization of soluble sarcoplasmic reticulum Ca‐ATPase

Vilsen

Andersen

1987

European Journal of Biochemistry

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The purpose of the present study was to elucidate the separate roles of lipid, detergent and protein-protein interaction for stability and catalytic properties of sarcoplasmic reticulum Ca-ATPase solubilized in the non-ionic detergent octa(ethy1ene glycol) monododecyl ether (Cl 2E8). The use of large-zone high-performance liquid chromatography permitted us to define the self-association state of Ca-ATPase peptide at various detergent, phospholipid and protein concentrations, and also during enzymatic turnover with ATP. Conditions were established for monomerization of Ca-ATPase in the presence of a high concentration of phospholipid relative to detergent. The lipid-saturated monomeric preparation was relatively resistant to inactivation in the absence of Ca", whereas delipidated enzyme in monomeric or in oligomeric form was prone to inactivation.Kinetics of phosphoenzyme turnover were examined in the presence and absence of Mg2 + . Dephosphorylation rates were sensitive to Mg2', irrespective of whether the peptide was present in soluble monomeric form or was membrane-bound. C1 2E8-solubilized monomer without added phospholipid was, however, characterized by a fast initial phase of dephosphorylation in the absence of Mg2+. This was not observed with monomer saturated with phospholipid or with monomer solubilized in myristoylglycerophosphocholine or deoxycholate. The mechanism underlying this difference was shown to be a C1 2E8-induced acceleration of conversion of ADP-sensitive phosphoenzyme (Ell') to ADP-insensitive phosphoenzyme (EzP).The phosphoenzyme isomerization rate was also found to be enhanced by low-affinity binding of ATP, This was demonstrated both in membrane-bound and in soluble monomeric Ca-ATPase.Our results indicate that a single peptide chain constitutes the target for modulation of phosphoenzyme turnover by Mg2+ and ATP, and that detergent effects, distinct from those arising from disruption of proteinprotein contacts, are the major determinants of kinetic differences between C1 2E8-solubilized and membranebound enzyme preparations.

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Section: Discussionsupporting

confidence: 67%

Effect of phospholipid, detergent and protein‐protein interaction on stability and phosphoenzyme isomerization of soluble sarcoplasmic reticulum Ca‐ATPase

Vilsen

Andersen

1987

European Journal of Biochemistry

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“…Although the monomeric Ca-pump protein in C12E8 can hydrolyze ATP at essentially the same rate as the membranebound protein, significant differences between the two preparations are well established, mostly by the work of Moller and co-workers (3). Of particular relevance to the present investigation is a difference in structural stability.…”

Section: Resultsmentioning

confidence: 87%

“…The sarcoplasmic reticulum (SR) Ca-pump protein can be solubilized in many nonionic detergents with full retention of ATPase activity (1)(2)(3). The detergent used in most previous work is dodecyl octaethyleneglycol monoether (C12E8), and this is the detergent used for the present investigation.…”

mentioning

confidence: 99%

Monomeric solubilized sarcoplasmic reticulum Ca pump protein: demonstration of Ca binding and dissociation coupled to ATP hydrolysis.

Martin¹,

Tanford²,

Reynolds³

1984

Proc. Natl. Acad. Sci. U.S.A.

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The sarcoplasmic reticulum Ca-pump protein can be solubilized in monomeric form in nonionic detergents with full retention of ATPase activity. It is impossible to prove directly that coupling between ATP hydrolysis and Ca2+ transport is maintained in the soluble state, because separate compartments for Ca21 uptake and Ca2' discharge are required to demonstrate this, but here we provide strong indirect evidence that coupling in fact persists, in both the forward and reverse directions of the normal pump cycle. Demonstration of coupling in the forward direction makes use of the fact that the solubilized protein is structurally labile in the absence of bound Ca2+. Loss of activity accompanying ATP hydrolysis in solution is quantitatively consistent with sequential Ca21 binding and dissociation during the hydrolysis cycle. Coupling in the reverse direction is demonstrated by the dependence of ATP synthesis on Ca2+ concentration. Although substantial differences between solubilized (monomeric) and membranebound protein are shown to exist, as previously reported, they do not affect the main conclusion from this work, which is that the molecular machinery for free-energy coupling in active Ca21 transport is an inherent property of each individual catalytic polypeptide chain of the pump protein.

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“…Ca2' translocation is dependent on conformation changes in this protein (1,2), but details of the translocation mechanism remain largely unresolved. The Ca2+-ATPase polypeptide is large (Mr = 115,000), traverses the lipid bilayer several times (3), and may provide the full structural basis for the translocation process (4). On the other hand, structural studies suggest that the Ca2+-ATPase peptides may form dimeric or higher oligomeric complexes in the membrane, which could be the minimal transport units (5)(6)(7).…”

mentioning

confidence: 99%

“…On the other hand, structural studies suggest that the Ca2+-ATPase peptides may form dimeric or higher oligomeric complexes in the membrane, which could be the minimal transport units (5)(6)(7). In favor of the view that the functional unit in Ca2+ transport is a monomeric peptide (8) are findings that show that it is possible to prepare soluble monomeric Ca2+-ATPase in detergent with retention of important functional properties, such as ATP hydrolysis (3) accompanied by release of Ca2+ from the two high-affinity transport sites (9,10) and ability to catalyze resynthesis of ATP from ADP and Pi after a "jump" in Ca2+ concentration (10)(11)(12). However, based on kinetic evidence obtained with soluble preparations of oligomeric and monomeric Ca2+-ATPase, it has been argued that peptide-peptide interactions are required for coupling between the ATPase reaction and Ca2+ translocation (13)(14)(15).…”

mentioning

confidence: 99%

Direct demonstration of structural changes in soluble, monomeric Ca2+-ATPase associated with Ca2+ release during the transport cycle.

Andersen¹,

Jørgensen²,

Møller³

1985

Proc. Natl. Acad. Sci. U.S.A.

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The time courses of changes in protein conformation and Ca2' binding in the phosphorylated state of membrane-bound and soluble monomeric Ca2+-ATPase from sarcoplasmic reticulum have been examined at pH 8.0, 20C. respectively. E1-P represents ADP-sensitive phosphoenzyme (highenergy acylphosphate) and E2-P represents ADP-insensitive phosphoenzyme (low-energy acylphosphate). E1-P and E2-P have high-and low-affinity Ca2' binding sites, respectively. transport sites, resulting in reorientation of the latter and transfer of free energy from the acylphosphate to Ca2+.The present communication deals with the possibility of demonstrating such conformational changes in the soluble monomeric Ca2+-ATPase peptide. The minimal scheme for the transport cycle shown as Fig. 1 provides the basis for design and discussion of the experiments. This scheme, which is based on studies of membrane-bound Ca2+-ATPase (2), implies that only two major conformational states of the protein (E1 and E2) are involved in Ca2+ translocation and that energy transfer is a single-step process occurring in relation to the E1P --E2P transition (1) (EP = phosphorylated Ca2+-ATPase). Our data support this view and show that the scheme of Fig. 1 4573The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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The sarcoplasmic reticulum Ca2+-ATPase

Cited by 113 publications

References 185 publications

Effect of phospholipid, detergent and protein‐protein interaction on stability and phosphoenzyme isomerization of soluble sarcoplasmic reticulum Ca‐ATPase

Effect of phospholipid, detergent and protein‐protein interaction on stability and phosphoenzyme isomerization of soluble sarcoplasmic reticulum Ca‐ATPase

Monomeric solubilized sarcoplasmic reticulum Ca pump protein: demonstration of Ca binding and dissociation coupled to ATP hydrolysis.

Direct demonstration of structural changes in soluble, monomeric Ca2+-ATPase associated with Ca2+ release during the transport cycle.

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