Common structural domains in the sarcoplasmic reticulum Ca-ATPase and the transverse tubule Mg-ATPase.

Damiani, Ernesto; Margreth, Alfredo; Furlan, Andrei; Dahms, A. Stephen; Arnn, J; Sabbadini, Roger A.

doi:10.1083/jcb.104.3.461

Cited by 27 publications

(7 citation statements)

References 46 publications

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“…1, lane 1),. in agreement with earlier observations on isolated TT from chicken muscle using the same procedure (Damiani et al, 1987). Junctional TT were also distinguishable on the basis of a more heterogeneous peptide Fig.…”

Section: Protein Composition Of Tt Membrane Fractions and Extent Of Csupporting

confidence: 92%

Co-localization of the dihydropyridine receptor and the cyclic AMP-binding subunit of an intrinsic protein kinase to the junctional membrane of the transverse tubules of skeletal muscle

Salvatori¹,

Damiani²,

Barhanin

et al. 1990

Biochemical Journal

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Junctional transverse tubules (TT) isolated from triads of rabbit skeletal muscle by centrifugation in an ion-free sucrose gradient were compared with membrane subfractions, predominantly derived from the free portion of TT, that had been purified from sarcoplasmic reticulum membrane contaminants by three different methods. The markers used were diagnostic membrane markers and the dihydropyridine (DHP) receptor, which is a specific marker of the junctional membrane of TT. Junctional TT have a high membrane density (Bmax. 60 pmol/mg of protein) of high-affinity (Kd 0.25 nM) DHP-binding sites using [3H]PN200-110 as the specific ligand. When analysed by SDS/PAGE under reducing conditions and by Western blot techniques, the TT were found to contain a concanavalin A-binding 150 kDa glycoprotein which probably corresponds to the alpha 2-subunit of the DHP receptor. This conclusion was supported by correlative immunoblot experiments with a specific antibody. Junctional TT are further distinguished from free TT by the presence of a high number (Bmax. 20 pmol/mg of protein) of [3H]cyclic AMP receptor sites, as determined by the Millipore filtration technique of Gill & Walton [(1974) Methods Enzymol. 38, 376-381]. Use of this method means that the number of receptors may have been underestimated. The TT-bound cyclic AMP receptor was identified as a 55 kDa protein by specific photoaffinity labelling with 8-N3-[3H]cyclic AMP, and had similar phosphorylation properties and apparent molecular mass to the RII form of the regulatory subunit of cyclic AMP-dependent protein kinase. Co-localization of the intrinsic cyclic AMP-dependent protein kinase and of the DHP receptor complex to the junctional membrane of TT supports the hypothesis that the 170 kDa alpha 1-subunit of the receptor is a substrate for the kinase.

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Section: Protein Composition Of Tt Membrane Fractions and Extent Of Csupporting

confidence: 92%

Co-localization of the dihydropyridine receptor and the cyclic AMP-binding subunit of an intrinsic protein kinase to the junctional membrane of the transverse tubules of skeletal muscle

Salvatori¹,

Damiani²,

Barhanin

et al. 1990

Biochemical Journal

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“…The approx. 100 kDa protein component of this enzyme, contrary to a previous report [1], was demonstrated to be structurally distinct from the sarcoplasmic-reticulum Ca2+-ATPase [2]. This finding is consistent with the well-established enzymic differences between the two enzymes.…”

Section: Introductioncontrasting

confidence: 54%

The Mg2+-ATPase of rabbit skeletal-muscle transverse tubule is a highly glycosylated multiple-subunit enzyme

Kirley

1991

Biochemical Journal

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The Mg2+-ATPase present in rabbit skeletal-muscle transverse tubules is an integral membrane enzyme which has been solubilized and purified previously in this laboratory [Kirley (1988) J. Biol. Chem. 263, 12682-12689]. The present study indicates that, in addition to the approx. 100 kDa protein (distinct from the sarcoplasmic-reticulum Ca2+-ATPase) seen previously to co-purify with the Mg2+-ATPase activity, there are also proteins having molecular masses of 160, 70 and 43 kDa. The 70 and 43 kDa glycosylated proteins (50 and 31 kDa after deglycosylation) are difficult to detect by SDS/PAGE before deglycosylation, owing to the broadness of the bands. Additional purification procedures, crosslinking studies and chemical and enzymic deglycosylation studies were undertaken to determine the structure and relationship of these proteins. Both the 97 and 160 kDa proteins were demonstrated to be N-glycosylated at multiple sites, the 97 kDa protein being reduced to a peptide core of 84 kDa and the 160 kDa protein to a peptide core of 131 kDa after deglycosylation. Although the Mg2+-ATPase activity is resistant to a number of chemical modification reagents, crosslinking inactivates the enzyme at low concentrations. This inactivation is accompanied by cross-linking of two 97 kDa molecules to one another, suggesting that the 97 kDa protein is involved in ATP hydrolysis. The existence of several proteins along with the inhibition of ATPase activity by cross-linking is consistent with the interpretation of the susceptibility of this enzyme to inactivation by most detergents as being due to the disruption of a protein complex of associated subunits by the inactivating detergents. The 160 kDa glycoprotein can be partially resolved from the Mg2+-ATPase activity, and is identified by its N-terminal amino acid sequence as angiotensin-converting enzyme.

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“…Although there was a report that the ecto-ATPase from rat liver had been cloned and sequenced (37), it seems likely that the Cell-CAM-105 adhesion protein that was cloned is not the rat liver ecto-ATPase (contrary to what was previously believed), since it can be partially separated chromatographically from the Cell-CAM-105 adhesion protein. 3 Therefore, the protein that was cloned and sequenced and used as an antigen for the production of antibodies was not the ecto-ATPase, but instead it was the co-purifying Cell-CAM-105 adhesion molecule (40). By analogy with both the rabbit skeletal muscle and the chicken smooth muscle ecto-ATPases that have been purified to homogeneity in our laboratory (which are both approximately 70-kDa proteins (8,18)), the rat liver ecto-ATPase is probably the approximately 70-kDa protein first identified as the ecto-ATPase (41).…”

Section: Discussionmentioning

confidence: 99%

“…1 The abbreviations used are: ecto-ATPase, extracellular adenosine that exhibit divalent cation-dependent NTPase activity on the extracellular side of the plasma membrane. These enzymes are present in tissues in low abundance and have low substrate specificity but have a very high specific activity (1)(2)(3)(4)(5)(6)(7)(8)(9). The identities and functions of ecto-ATPases are the subject of a recent review in which the nomenclature of "E-type ATPases" was proposed to describe these enzymes (10).…”

Section: Properties Of and Proteins Associated With The Extracellular Atpase Of Chicken Gizzard Smooth Musclementioning

confidence: 99%

Properties of and Proteins Associated with the Extracellular ATPase of Chicken Gizzard Smooth Muscle

Stout

Strobel

Kirley

1995

Journal of Biological Chemistry

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The chicken gizzard smooth muscle extracellular ATPase (ecto-ATPase) is a low abundance, high specific activity, divalent cation-dependent, nonspecific nucleotide triphosphatase (NTPase). The ATPase is a 66-kDa glycoprotein with a protein core of 53 kDa (Stout, J.G. and Kirley, T.L. (1994) J. Biochem. Biophys. Methods 29, 61-75). In this study we evaluated the characteristics of a bank of monoclonal antibodies raised against a partially purified chicken gizzard ecto-ATPase. 18 monoclonal antibodies identified by an ATPase capture assay were tested for effects on ATPase activity as well as for their Western blot and immunoprecipitation potential. The five most promising monoclonal antibodies were used to immunopurify the ecto-ATPase. The one-step immunoaffinity purification of solubilized chicken gizzard membranes with all five of these monoclonal antibodies isolated a 66-kDa protein whose identity was confirmed by N-terminal sequence analysis to be the ecto-ATPase. Several of these monoclonal antibodies stimulated ecto-ATPase activity similar to that observed previously with lectins. Western blot analysis revealed that three of the five monoclonal antibodies recognized a major immunoreactive band at 66 kDa (53-kDa core protein), consistent with previous purification results. The other two antibodies recognized proteins of approximately 90 and 160 kDa on Western blots. The 90-kDa co-immunopurifying (and presumably associated or related) protein was identified by N-terminal analysis as LEP100, a glycoprotein that shuttles between the plasma and lysosomal membranes. The approximately 160-kDa co-immunopurifying protein was identified by N-terminal analysis as integrin, a protein involved in extracellular contacts with adhesion molecules. Extended N-terminal sequence analysis of the immunopurified 66-kDa ecto-ATPase revealed some sequence homology with mouse lysosomal associated membrane protein. Tissue distribution of the ecto-ATPase showed that the highest levels of protein were expressed in muscle tissues (cardiac, skeletal, and smooth) and brain.

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Common structural domains in the sarcoplasmic reticulum Ca-ATPase and the transverse tubule Mg-ATPase.

Cited by 27 publications

References 46 publications

Co-localization of the dihydropyridine receptor and the cyclic AMP-binding subunit of an intrinsic protein kinase to the junctional membrane of the transverse tubules of skeletal muscle

Co-localization of the dihydropyridine receptor and the cyclic AMP-binding subunit of an intrinsic protein kinase to the junctional membrane of the transverse tubules of skeletal muscle

The Mg2+-ATPase of rabbit skeletal-muscle transverse tubule is a highly glycosylated multiple-subunit enzyme

Properties of and Proteins Associated with the Extracellular ATPase of Chicken Gizzard Smooth Muscle

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