Characterization of a novel alpha-tocopherol-binding-protein from bovine heart cytosol

Margaret J. Gordon, Fiona M. Campbell, Garry G. Duthie, Asim K. Dutta-Roy

Research output: Contribution to journalArticle

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Abstract

We previously reported the identification of a new alpha-tocopherol-binding protein (similar to 15 kDa) in the cytosol of rat liver and heart and in rabbit heart (A, K, Dutta-Roy et al., J, Nutr, Biochem, 5, 562-570, 1994). This protein specifically binds cr-tocopherol and enhances its transfer between separate membranes, In the present paper we have purified and characterized the alpha-tocopherol-binding protein from bovine heart cytosol and compared its various structural and functional properties with the similar size (similar to 15 kDa) cytosolic fatty acid-binding protein of this tissue, alpha-Tocopherol-binding protein was purified to electrophoretic homogeneity from bovine heart cytosol by a procedure involving precipitation with 70% ammonium sulfate, followed sequentially by gel filtration chromatography and chromatofocusing, The purified protein migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 16 kDa, Isoelectric focusing of the purified protein showed that the pI value is around 4.5, The binding of alpha-tocopherol to the purified protein was rapid, reversible, and saturable, The alpha-tocopherol-binding protein did not bind oleate as assessed by direct radiolabeled fatty acid binding and fluorescence enhancement assay, Amino acid analysis showed the presence of a large number of Ala, Gly, Ser, Lys, and Pro residues and a lesser number of aromatic residues in this protein, Anti-bovine heart fatty acid-binding protein antibody did not recognize the alpha-tocopherol-binding protein in the Western blot, The Western blot, ligand affinity, molecular size, and amino acid analysis data suggest that the alpha-tocopherol-binding protein is different from the cytosolic fatty acid-binding protein and that it may be involved in intracellular transport and metabolism of alpha-tocopherol. (C) 1995 Academic Press,Inc.

Original languageEnglish
Pages (from-to)140-146
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume318
Issue number1
DOIs
Publication statusPublished - 1 Apr 1995

Keywords

  • alpha-tocopherol binding protein
  • bovine heart
  • alpha-tocopherol
  • amino-acid analysis
  • fatty acid-binding protein
  • vitamin-e
  • sterol carrier protein-2
  • rat-liver
  • purification
  • identification
  • distinct
  • tissue

Cite this

Characterization of a novel alpha-tocopherol-binding-protein from bovine heart cytosol. / Gordon, Margaret J.; Campbell, Fiona M.; Duthie, Garry G.; Dutta-Roy, Asim K.

In: Archives of Biochemistry and Biophysics, Vol. 318, No. 1, 01.04.1995, p. 140-146.

Research output: Contribution to journalArticle

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abstract = "We previously reported the identification of a new alpha-tocopherol-binding protein (similar to 15 kDa) in the cytosol of rat liver and heart and in rabbit heart (A, K, Dutta-Roy et al., J, Nutr, Biochem, 5, 562-570, 1994). This protein specifically binds cr-tocopherol and enhances its transfer between separate membranes, In the present paper we have purified and characterized the alpha-tocopherol-binding protein from bovine heart cytosol and compared its various structural and functional properties with the similar size (similar to 15 kDa) cytosolic fatty acid-binding protein of this tissue, alpha-Tocopherol-binding protein was purified to electrophoretic homogeneity from bovine heart cytosol by a procedure involving precipitation with 70{\%} ammonium sulfate, followed sequentially by gel filtration chromatography and chromatofocusing, The purified protein migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 16 kDa, Isoelectric focusing of the purified protein showed that the pI value is around 4.5, The binding of alpha-tocopherol to the purified protein was rapid, reversible, and saturable, The alpha-tocopherol-binding protein did not bind oleate as assessed by direct radiolabeled fatty acid binding and fluorescence enhancement assay, Amino acid analysis showed the presence of a large number of Ala, Gly, Ser, Lys, and Pro residues and a lesser number of aromatic residues in this protein, Anti-bovine heart fatty acid-binding protein antibody did not recognize the alpha-tocopherol-binding protein in the Western blot, The Western blot, ligand affinity, molecular size, and amino acid analysis data suggest that the alpha-tocopherol-binding protein is different from the cytosolic fatty acid-binding protein and that it may be involved in intracellular transport and metabolism of alpha-tocopherol. (C) 1995 Academic Press,Inc.",
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AU - Campbell, Fiona M.

AU - Duthie, Garry G.

AU - Dutta-Roy, Asim K.

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N2 - We previously reported the identification of a new alpha-tocopherol-binding protein (similar to 15 kDa) in the cytosol of rat liver and heart and in rabbit heart (A, K, Dutta-Roy et al., J, Nutr, Biochem, 5, 562-570, 1994). This protein specifically binds cr-tocopherol and enhances its transfer between separate membranes, In the present paper we have purified and characterized the alpha-tocopherol-binding protein from bovine heart cytosol and compared its various structural and functional properties with the similar size (similar to 15 kDa) cytosolic fatty acid-binding protein of this tissue, alpha-Tocopherol-binding protein was purified to electrophoretic homogeneity from bovine heart cytosol by a procedure involving precipitation with 70% ammonium sulfate, followed sequentially by gel filtration chromatography and chromatofocusing, The purified protein migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 16 kDa, Isoelectric focusing of the purified protein showed that the pI value is around 4.5, The binding of alpha-tocopherol to the purified protein was rapid, reversible, and saturable, The alpha-tocopherol-binding protein did not bind oleate as assessed by direct radiolabeled fatty acid binding and fluorescence enhancement assay, Amino acid analysis showed the presence of a large number of Ala, Gly, Ser, Lys, and Pro residues and a lesser number of aromatic residues in this protein, Anti-bovine heart fatty acid-binding protein antibody did not recognize the alpha-tocopherol-binding protein in the Western blot, The Western blot, ligand affinity, molecular size, and amino acid analysis data suggest that the alpha-tocopherol-binding protein is different from the cytosolic fatty acid-binding protein and that it may be involved in intracellular transport and metabolism of alpha-tocopherol. (C) 1995 Academic Press,Inc.

AB - We previously reported the identification of a new alpha-tocopherol-binding protein (similar to 15 kDa) in the cytosol of rat liver and heart and in rabbit heart (A, K, Dutta-Roy et al., J, Nutr, Biochem, 5, 562-570, 1994). This protein specifically binds cr-tocopherol and enhances its transfer between separate membranes, In the present paper we have purified and characterized the alpha-tocopherol-binding protein from bovine heart cytosol and compared its various structural and functional properties with the similar size (similar to 15 kDa) cytosolic fatty acid-binding protein of this tissue, alpha-Tocopherol-binding protein was purified to electrophoretic homogeneity from bovine heart cytosol by a procedure involving precipitation with 70% ammonium sulfate, followed sequentially by gel filtration chromatography and chromatofocusing, The purified protein migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 16 kDa, Isoelectric focusing of the purified protein showed that the pI value is around 4.5, The binding of alpha-tocopherol to the purified protein was rapid, reversible, and saturable, The alpha-tocopherol-binding protein did not bind oleate as assessed by direct radiolabeled fatty acid binding and fluorescence enhancement assay, Amino acid analysis showed the presence of a large number of Ala, Gly, Ser, Lys, and Pro residues and a lesser number of aromatic residues in this protein, Anti-bovine heart fatty acid-binding protein antibody did not recognize the alpha-tocopherol-binding protein in the Western blot, The Western blot, ligand affinity, molecular size, and amino acid analysis data suggest that the alpha-tocopherol-binding protein is different from the cytosolic fatty acid-binding protein and that it may be involved in intracellular transport and metabolism of alpha-tocopherol. (C) 1995 Academic Press,Inc.

KW - alpha-tocopherol binding protein

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KW - alpha-tocopherol

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KW - vitamin-e

KW - sterol carrier protein-2

KW - rat-liver

KW - purification

KW - identification

KW - distinct

KW - tissue

U2 - 10.1006/abbi.1995.1214

DO - 10.1006/abbi.1995.1214

M3 - Article

VL - 318

SP - 140

EP - 146

JO - Archives of Biochemistry and Biophysics

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ER -