Purification of Morphologically Intact Triad Structures from Skeletal Muscle

Overview

Journal J Cell Biol

Specialty Cell Biology

Date 1983 Apr 1

PMID 6300142

Citations 37

Authors

R D Mitchell

P Palade

S Fleischer

Affiliations

Soon will be listed here.

Abstract

A procedure has been devised for isolation of triads (t-tubule/sarcoplasmic reticulum (SR) junctional complexes) from rabbit skeletal muscle. The procedure consists of preparation of a heavy microsomal fraction followed by two sequential 90-min sucrose gradient centrifugations to enrich the triads. A pyrophosphate/phosphate/magnesium buffer system was introduced to decrease aggregation in order to achieve effective separation. The preparation time is 12 h. Some differences between purified triads isolated by two variants of this method are noted. The purity of the triad fractions has been estimated by particle counting to be in the vicinity of 50%. There is good retention of morphology and Ca++-loading activity and enrichment in Na+,K+-ATPase and adenylate cyclase. The triads are practically devoid of contractile elements, mitochondria, and free plasmalemma, and low in content of light SR. The method for obtaining enriched triads is reproducible, and sufficient yields are obtained for structural, biochemical, and functional characterization.

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References

Caswell A, Lau Y, Garcia M, Brunschwig J . Recognition and junction formation by isolated transverse tubules and terminal cisternae of skeletal muscle. J Biol Chem. 1979; 254(1):202-8. View

Walseth T, Johnson R . The enzymatic preparation of [alpha-(32)P]nucleoside triphosphates, cyclic [32P] AMP, and cyclic [32P] GMP. Biochim Biophys Acta. 1979; 562(1):11-31. DOI: 10.1016/0005-2787(79)90122-9. View

Jones L, Besch Jr H, Fleming J, McConnaughey M, Watanabe A . Separation of vesicles of cardiac sarcolemma from vesicles of cardiac sarcoplasmic reticulum. Comparative biochemical analysis of component activities. J Biol Chem. 1979; 254(2):530-9. View

Franzini-Armstrong C . STUDIES OF THE TRIAD : I. Structure of the Junction in Frog Twitch Fibers. J Cell Biol. 2009; 47(2):488-99. PMC: 2108094. DOI: 10.1083/jcb.47.2.488. View

Bers D . Isolation and characterization of cardiac sarcolemma. Biochim Biophys Acta. 1979; 555(1):131-46. DOI: 10.1016/0005-2736(79)90078-6. View

Barchi R, Weigele J, Chalikian D, Murphy L . Muscle surface membranes: preparative methods affect apparent chemical properties and neurotoxin binding. Biochim Biophys Acta. 1979; 550(1):59-76. DOI: 10.1016/0005-2736(79)90115-9. View

Colaco C, Evans W . Partial purification of an intercalated disc-containing cardiac plasma membrane fraction. Biochim Biophys Acta. 1982; 684(1):40-6. DOI: 10.1016/0005-2736(82)90046-3. View

Meissner G, Conner G, Fleischer S . Isolation of sarcoplasmic reticulum by zonal centrifugation and purification of Ca 2+ -pump and Ca 2+ -binding proteins. Biochim Biophys Acta. 1973; 298(2):246-69. DOI: 10.1016/0005-2736(73)90355-6. View

Scarpa A . Measurements of cation transport with metallochromic indicators. Methods Enzymol. 1979; 56:301-38. DOI: 10.1016/0076-6879(79)56030-3. View

10.

Lau Y, Caswell A, Brunschwig J . Isolation of transverse tubules by fractionation of triad junctions of skeletal muscle. J Biol Chem. 1977; 252(15):5565-74. View

11.

Endo M . Calcium release from the sarcoplasmic reticulum. Physiol Rev. 1977; 57(1):71-108. DOI: 10.1152/physrev.1977.57.1.71. View

12.

Fleischer S, Brierley G, KLOUWEN H, SLAUTTERBACK D . Studies of the electron transfer system. 47. The role of phospholipids in electron transfer. J Biol Chem. 1962; 237:3264-72. View

13.

Brunschwig J, Brandt N, Caswell A, Lukeman D . Ultrastructural observations of isolated intact and fragmented junctions of skeletal muscle by use of tannic acid mordanting. J Cell Biol. 1982; 93(3):533-42. PMC: 2112139. DOI: 10.1083/jcb.93.3.533. View

14.

Lau Y, Caswell A, Garcia M, Letellier L . Ouabain binding and coupled sodium, potassium, and chloride transport in isolated transverse tubules of skeletal muscle. J Gen Physiol. 1979; 74(3):335-49. PMC: 2228522. DOI: 10.1085/jgp.74.3.335. View

15.

Caswell A, Baker S, Boyd H, Potter L, Garcia M . beta-adrenergic receptor and adenylate cyclase in transverse tubules of skeletal muscle. J Biol Chem. 1978; 253(9):3049-54. View

16.

Kono T, COLOWICK S . Isolation of skeletal muscle cell membrane and some of its properties. Arch Biochem Biophys. 1961; 93:520-33. DOI: 10.1016/s0003-9861(61)80046-5. View

17.

Cadwell J, Caswell A . Identification of a constituent of the junctional feet linking terminal cisternae to transverse tubules in skeletal muscle. J Cell Biol. 1982; 93(3):543-50. PMC: 2112169. DOI: 10.1083/jcb.93.3.543. View

18.

Seiler S, Fleischer S . Isolation of plasma membrane vesicles from rabbit skeletal muscle and their use in ion transport studies. J Biol Chem. 1982; 257(22):13862-71. View

19.

LOWRY O, ROSEBROUGH N, FARR A, RANDALL R . Protein measurement with the Folin phenol reagent. J Biol Chem. 1951; 193(1):265-75. View

20.

Caswell A, Lau Y, Brunschwig J . Ouabain-binding vesicles from skeletal muscle. Arch Biochem Biophys. 1976; 176(2):417-30. DOI: 10.1016/0003-9861(76)90184-3. View